__________________________________________________________________________________________________

Preface: (char lhungzt)

Hello! Soo this is my compilation on biochemistry. It’s quite long and over-info’d but then again there’s nothing wrong
with a little extra information . I have no clue if any of the topics here will come out during discussions and lessons
(hopefully not all) but when it does, you’ll be ready! (charrr)
There’s no guarantee this whole compilation is accurate, if you see any errors please inform me so we can change it
and update it
The Color Content is very monotonous (I really really really like the color yellow on my notes hehehe)
Lastly, have fun studying! (yeah, I know that’s kind of a stretch but hey you gotta motivate yourself)

References:
Lippincotts Illustrated Reviews: Biochemistry, 5th Edition – by Ferrier, Denise R. (2011)
Biochemistry (Velez College)

–everything that you’ve come to expect–

BIOCHEMISTRY
– study of the chemical and physicochemical process occurring within a living organism
– study of the compounds, chemical reactions, and molecular interactions that are involved in the production,
maintenance, and reproduction of living organisms.

CHEMICAL COMPOSITION OF LIVING MATTER

• Tissues are made up of about 70-90% water & 10-30% solids. Only 1% of the solids are inorganic & the rest is organic
• The water component is found partly in the free state, that is, capable of passing from one cell to another. This passage
is governed by the physical laws of osmosis and diffusion.
↳ Any disturbance in the osmotic equilibrium leads to an alteration in the water distribution.
↳ The other portion of water is fixed, or held in a state of hydrophilic colloid system, thereby forming a constant
constituent of the cell itself. This condition is responsible for the maintenance of the shape and size of the cells
↳ The amount of this fixed water in the tissues varies with their functional activities
• Tissues with greater physiologic activity have larger water content than those with less activity
• Tissues like blood, brain, and muscles have high water content while protective and supporting tissues like bones,
teeth and adipose tissues have low water content
↳ The cartilage, however, is an exception, because although it is inert, it has a high water content
• The Inorganic Substances are made up of Sodium, Potassium, Calcium, Magnesium, and Ammonium in the form of
Chlorides, Sulfates, Phosphates, Carbonates, etc
↳ Traces of Iron, Iodine, Copper, Manganese, Cobalt, Zinc are also present in combination with organic radicals
↳ The rest of the protoplasm is made up of organic compounds such as carbohydrates, lipids, and proteins

1
HIERARCHY OF BIOLOGICAL ORGANIZATION
Organism
Organs
Tissues
Cells
Nucleus Mitochondria Chloroplasts
Multi-Enzyme Complexes Ribosomes Chromosomes Membranes Structural Elements Contractile Systems
Proteins Nucleic Acids Polysaccharide Phospholipids
↳Amino Acid ↳Nucleotides ↳Sugars ↳Fatty Acid
CO2 and H2O
* Start from the bottom going up. All stem from the one before it except the ones shaded with gray . All shaded gray stops there and doesn’t
proceed upward, only the Nucleic Acid progresses forwards.

BUILDING BLOCKS OF LIFE SIMPLE MOLECULES

1 Proteins 1 • Elements of the biosphere – the fundamental building blocks
2 Carbohydrates • Major Elements (4)
3 Lipids Carbon – Hydrogen – Oxygen – Nitrogen | CHON
4 Nucleic Acids – 4 most abundant elements

Carbon – forms the basis of all organic molecules. Has the ability to form long chains by electron sharing – in
C
which chains maybe linear, branched, cyclic, or a combination so that very large structures are made possible
Oxygen-Hydrogen-Nitrogen – readily forms strong covalent bonds with Carbon and with each other, thus
OHN
increasing the possible number, shape, and size of the molecules
• Minor Elements (7)
Calcium – Phosphorus – Potassium – Sulfur – Chlorine – Magnesium – Sodium | Ca-P-K-S-Cl-Mg-Na
Calcium & • The biggest constituents of the minor elements
Phosphorus • In man, most of the calcium and phosphorus are part of the structure of the bones
• Plays a key role in the regulation of cellular processes
Calcium
• Affects the contraction of the heart and skeletal muscles
• Important part of the phospholipid structure which form the basis of membranes
Phosphorus • Vital part of compounds involved in energy transfer such as ATP which is often described as the
energy currency of the cell
• Adequate amount of these elements must be supplied in the diet to maintain life
Mg-Na-S-K-Cl • Na+ ions are the major cations of the extracellular fluid (blood plasma)
• K+ ions are predominant in the intracellular fluid (liver cell)
• Trace and Ultra-trace Elements – play a key role in maintaining life
Iron (Fe) – needed for O2 transport Iodine (I) – essential part of the thyroid hormone that controls metabolic rate
2 Water – the liquid environment of living organisms • Molecule which is absolutely essential for life
3 Gases of the Atmosphere – interaction with biosphere • From the 4 major elements (CO2, H2O Vapor, O2, N2)

SMALL ORGANIC MOLECULES

1 Amino Acids – the building block of proteins 4 Phospholipids – structural elements of membranes
2 Monosaccharides – basic unit of carbohydrates 5 Steroids – bile salts and hormones
3 Triacylglycerides – long-term energy stores

MACROMOLECULES
1 Proteins – catalysts, structural elements, chemical messengers, transport agents, protective barriers
2 Carbohydrates – fuel, structural elements, lubricants, cellular communication agents
3 Lipids – fuel, structural elements, membrane components, transport agents, chemical messengers
4 DNA – storage of genetic information RNA – catalysts, expression of Genetic Information

2
INTRODUCTION TO THE CELL
Prokaryotes vs Eukaryotes
Characteristic Prokaryotic Eukaryotic

Structure

Size of Cell 0.2-2.0 μm in diameter 10-100 μm in diameter

Nucleus No Nuclear Membrane or Nucleoli True Nucleus – Nuclear Membrane & Nucleoli
Membrane-Enclosed Absent Present: Ex. Includes Lysosomes, Golgi
Organelles Complex, ER, Mitochondria and Chloroplasts
Flagella Consists of two protein building blocks Complex – consists of microtubules
Glycocalyx Present – as a capsule or slime layer Present in some cells that lack a cell wall
Usually Present – Chemically Complex When Present – Chemically Simple (includes
Cell Wall
(bacterial cell wall includes peptidoglycan) cellulose and chitin)
No Plasma Membrane No Carbohydrates – generally lack sterols Sterols & Carbohydrates – serve as receptors
Cytoplasm No Cytoskeleton or Cytoplasmic Streaming Cytoskeleton – Cytoplasmic Streaming
Ribosomes Small (70S) Large (80S) and Small (70S) in organelles
Usually Singular Chromosome – typically Multiple Linear Chromosomes with Histones
Chromosomes (DNA)
lacks Histones
Cell Division Binary Fission Involves Mitosis
Sexual Recombination None – transfer of DNA only Involves Meiosis

CHEMICAL COMPOSITION OF THE CELL | Inorganic

Percentage
of Body Element Use
Weight (%)
65 Oxygen Found in blood/lungs due to respiration
18.6 Carbon Found in every organic molecule in body/ waste product of respiration
9.7 Hydrogen Found in water and various body tissues
3.2 Nitrogen Found in proteins and organic compounds, and in lungs due to abundance in atmosphere
1.8 Calcium Found in skeletal system, including teeth; nervous system, muscles, and blood
1.0 Phosphorus Found in bones, teeth, and nucleic acids
0.4 Potassium Found in muscles, nerves, and certain tissues
0.2 Sodium Found in muscles and nerves; excreted in sweat
0.2 Chlorine Found in skin to facilitate water absorption by cells
0.06 Magnesium Serves as cofactor for various enzymes in the body
0.04 Sulfur Found in amino acids and proteins
0.007 Iron Found mostly in blood since it facilitates oxygen transport
0.0002 Iodine Found in thyroid gland hormones
*For Inorganic Components

3
CHEMICAL COMPOSITION OF THE CELL | Organic
Carbohydrates Lipids Proteins Nucleic Acids
•Energy Supply and •Energy Storage and Source •Building Blocks of many •Contain Genetic
Storage •Reduces loss of water by Structural components of cell; Information of Cells
evaporation required for Growth
•Structural Support •Source of Metabolic Water •Form enzymes and Hormones •Vital role in Protein
to Cell Walls •Structural role in Cell Membrane Synthesis
*For Organic Components

STRUCTURAL PARTS OF A CELL

PARTS OF A CELL
Nucleus Cytoplasm
Chromatin Ribosomes
Chromosomes Endoplasmic Reticulum
Chromosomes Golgi Apparatus
Nucleoli Lysosomes
Nuclear Membrane Mitochondria
Plasma Membrane Centrioles

4
Cell Parts Description Function
Spherical or oval shaped structure that usually Contains genes, which control cellular structure
Nucleus
is the most prominent feature of a cell and direct cellular functions
Fine and unevenly stained network composed Provides a physical means for packing the very
of about 30% DNA, which is traditionally called long DNA molecules in a compact, orderly way
Chromatin our genetic material, about 60% globular *Fundamental units of chromatin: Nucleosomes
histone proteins, and about 10% RNA chains, (Nuclear Bodies)
newly formed or forming
Highly coiled and folded DNA molecule that is Its compactness avoids entanglement and
combined with protein molecules breakage of the delicate chromatin strands
Chromosomes during the movements that occur during cell
division
*Human Somatic Cells have 46 chromosomes --
23 inherited from each parent
Cluster of protein, DNA, and RNA, that is not Sites of RNA synthesis and assembly of rRNA and
Nucleoli
enclosed by a membrane proteins into ribosomal units
Double membrane; Both layers are lipid Separates the nucleus from the cytoplasm
Nuclear
bilayers similar to the plasma membrane/
Membrane
nuclear envelope
Fluid-Mosaic Lipid Bilayer (phospholipids, Protects cellular contents; makes contact with
cholesterol, and glycolipids) studded with other cells; contains channels, transporters,
Plasma
proteins; surrounds cytoplasm receptors, enzymes, cell-identity markers, and
Membrane
linker proteins; mediates the entry and exit of
substances
Consist of the cellular contents between that Site of all intracellular activities except those
Cytoplasm plasma membrane and nucleus – cytosol and occurring in the nucleus
organelles
Composed of two subunits containing Protein Synthesis
Ribosomes ribosomal RNA and proteins; may be free in
cytosol or attached to rough ER
Membranous network of flattened sacs or Rough: Synthesis of glycoprotein and
tubules phospholipids that are transferred to cellular
Endoplasmic organelles, inserted into the plasma membrane,
Reticulum or secreted during exocytosis
Smooth: Synthesis of fatty acids and steroids,
Inactivates or detoxifies drugs
Consists of 3-20 flattened membranous sacs Form glycoproteins, glycololipids, and
called cisternae; structurally and functionally lipoproteins
Golgi Apparatus
divided into entry (cis) face, medial cisternae,
and exit (trans) face
Vesicle formed from Golgi complex that Fuses with and digests contents of endosomes;
contains digestive enzymes pinocytic vesicles, and phagosomes transports
Lysosomes final products of digestion into cytosol; digests
worn-out organelles (autophagy), entire cells
(autolysis), and extracellular materials
Consists of outer and inner mitochondrial Site of aerobic cellular respiration reactions that
Mitochondria membranes, cristae, and matrix; new produce most of cell’s ATP
mitochondria form from preexisting ones
Small, barrel-shaped organelles oriented at Centrosomes: Best known for its generation of
Centrioles right angles to each other microtubules and its role of organizing the
mitotic spindle in cell division

5
PROTEINS
• Most abundant macromolecule found within the living cell
• These are organic compounds of high molecular weight, made up of alpha amino acids joined by peptide linkage
• Mulder (1839) gave the name proteins (greek word proteios – preeminence)
• Most important of all biological substances, being the fundamental constituents of the protoplasm of the cell
COMPOSITION
• Protein molecule is known to contain carbon, hydrogen, oxygen, nitrogen, sulfur, phosphorus, together with traces of
iron, copper, iodine, manganese, and zinc. (S-P-O-N-C-H—I-Fe-Cu-Zi-Mn)
• Chemical structure of the protein molecule is yet undetermined
• Molecular Weight ranging from 5,000-8,000,000. It possesses all the properties and characteristics of a colloid.
• Protein molecule is composed of hundreds/thousands of amino acids linked by peptide linkages (carboxyl group of
one amino acid linked with the amine group of the other)
• On hydrolysis, either by enzymes or by prolonged boiling with acids or bases, the protein molecule is broken down
into fragments of decreasing complexity such as proteoses, peptones, polypeptides, and finally α-amino acids
• The structure of the α-amino acid is represented by the formula where R stands for either a hydrogen, aliphatic,
aromatic, or a heterocyclic component
Proteoses → Peptones → Polypeptides → α-Amino Acids

Protein No. of A.A. No. of MWH DYNAMIC STATE OF PROTEINS

residues P.C.
Insulin 51 2 5,733
Myoglobin 153 1 16,890
Hemoglobin 574 4 64,500
Collagen 3,000 3 300,000
*A.A. – amino acid **P.C. – Polypeptide Chains

Amino Acid Formula:

CLASSIFICATION OF AMINO ACIDS

ALIPHATIC AMINO ACIDS
• Alaninen(ala) – mother of all amino acids • Valinen (val) – important for the nervous system to function
• Leucinen (leu) – associated with digestive enzymes • Isoleucinen (ile) – also associated with digestive enzymes
• Prolinen (pro) – called imino acid – also called “helix breaker” because it disrupts the α-helical config. of -COOH;
inhibits the formation of an α-helix. It also does not contain any primary amino group.
• Glycinen (gly) – simplest amino acid – also a major excitatory neurotransmitter
↳ Optically inactive due to the absence of α -asymmetric C-atom
↳ Used by the body in the detoxification of benzoic acid
Note: the defect on the metabolism of val, leu, and ile will accumulate in tissues perhaps because of the absence of
enzymes and that would result to a certain disease called Maple Syrup Urine
AROMATIC AMINO ACIDS
• Phenylalaninen (phe) – important in the utilization of Vit. C and production of thyroxine
↳ Some people lack the enzyme necessary to convert phe to tyr. The phe that they obtain from their diet is instead
diverted along an alternative metabolic pathway which produces Phenylpyruvic Acid
↳ Phenylpyruvic Acid – can cause mental retardation in infants who are deficient in the enzymes necessary to convert
phe to tyr. They are said to suffer phenylketonuria – this disease cannot be cured, but controlled by restricting the
dietary intake of phe (meat are rich in phe)
• Tyrptophann (trp) – important in utilization of B-vitamins and synthesis of neurotransmitter
6
↳ Contained in charred potion of broiled fish and meat, which is responsible for cancer diseases
↳ Needed in the formation of neurotransmitters
↳ Used as a food supplement by some individuals seeking relief from depression, insomnia, and certain other nervous
system disorders
• Tyrosinen (tyr) – synthesis of some thyroid hormone
SULFUR CONTAINING AMINO ACIDS
n
• Methionine (met) – initiates protein synthesis & is participated in the transmethylation reactions as a methyl donor
• Cysteinen (cys) – found abundantly in natural curly hair
HYDROXYL-CONTAINING AMINO ACIDS
• Serinen (ser) – component of phosphoprotein – phosphate group is attached to serine
• Threoninen (thr) – building tissues & utilization of nutrients
AMIDE-CONTAINING AMINO ACIDS
n
• Asparagine (asn) – important site for asparagine-linked glycosylation (N-glycosylation) within peptides
• Glutaminen (gln) – has a special role in intestinal health
ACIDIC AMINO ACIDS
• Aspartic Acid– (asp) – its salt is called aspartate – major excitatory neurotransmitter
• Glutamic Acid– (glu) – its salt is called glutamate -- major excitatory neurotransmitter
BASIC AMINO ACIDS
• Lysine+ (lys) – found in collagen and aids in assimilation of other amino acids. Contains ε-amino group
• Arginine+ (arg) – used for heart and blood vessel conditions including congestive heart failure (CHF)
• Histidine+ (his) – has significant buffering capacity at physiological pH – the only a.a having this property
↳ Decarboxylation of Histidine – involves loss of -COO-group as CO2 forms histamine
↳ Histamine – powerful vasodilator, normally present in tissue and formed in excessive amounts under traumatic shock.
It is also responsible for many of the symptoms associated with high fever and other allergies
*All with the exponents beside their names are their charges. n= neutral, += positive, and –= negative
**All shaded in blue are non-polar and all those in green are polar.

ESSENTIAL AMINO ACIDS

– Must be included in dietary requirements. Dietary proteins must contain adequate amounts of essential amino acids
Phenylalanine Valine Threonine Tryptophan Isoleucine
Methionine Histidine Arginine Leucine Lysine
• All 10 essential amino acids can be summed by the mnemonic PVT TIM HALL
• Arginine & Histidine – required in childhood during periods of rapid growth • Other 8 – essential throughout life
CHEMICAL PROPERTIES OF AMINO ACIDS
Due to – COOH group Due to – NH2 group
• Salt formation and titration • Acylation
• Benzoylation
• Methylation
• Formation of esters and amides • Reaction with Sanger’s Reagent (1-Fluoro-2,4-dinitrobenzene)/ DNFB
• Reaction with nitrous acid (HNO2)
• Oxidative Deamination – removal of – M+2 group
• Formation of amino acyl chlorides • Reaction with formaldehyde (CH2O)
• Reaction with Aromatic Aldehyde in Alkali
• Reaction with Ninhydrin (C9H6O4) (oxidative decarboxylation)
• Decarboxylation – important in allergy • Reaction with CO2
reactions • Chelation with metal ions
• Peptide bond formation

7
PEPTIDE FORMATION
There is a union of –COOH and –NH2 group and elimination of H2O No. of peptide likages = No. of amino acids – 1

PEPTIDE BOND

• Covalent Bond – has a partial double bond character due to π e– delocalization

• The C-N bond in the peptide linkage has a partial double bond charater that make it rigid and prevent the adjacent
groups from rotating freely

PROOF OF π e– DELOCALIZTION
1 Bond Length 2 Bond Angle
Peptide C-N Normal C-N Peptide Normal
1.32 A 1.47 A 122 109
CONSEQUENCES OF π e– DELOCALIZTION
1 Imparts bond stability • Less susceptible to hydrolysis • Increase bond stability
2 Cannot be easily rotated • Conformation
Cis – unstable and effect is steric hindrance Trans – stable

8
PROTEINS
Amino Acids as the Building Block of Proteins
Proteins – are made up of amino acids. Amino acids are therefore “building blocks of proteins”
Consists of an amino group, a carboxyl group, a hydrogen atom and a distinctive R group bonded
to a carbon which is called the α-carbon. An R group is also called a side chain

Amino Acids

BASIC STRUCTURE OF AMINO ACIDS

• H2N-CH(R)-COOH • 20 α-amino acids found in proteins • All are primary amines except for Proline (secondary)
• Not all amino acids are alpha amino acids, e.g. β-Alanine is a naturally occurring β-amino acid
• Other exceptions include:
↳Sarcosine: N-Methyl Glycine – secondary amine ↳Betaine: N,N,N-Trimethylglycine – quaternary amine
• Most amino acids have a Carboxylic Acid (Carboxylate) as the acid functionality except for the amino acid taurine.
Taurine has a Sulfonic Acid
STEREOCHEMISTRY OF AMINO ACIDS
• The α-carbon of each amino acid is attached to four different chemical groups and is therefore a chiral or optically
active carbon atom.
• Amino acids that have an asymmetric center at the α-carbon can exist in two forms. Designated as D- and L- that are
mirror images of each other.
• The two forms in each pair are termed stereoisomers, optical isomers or enantiomers.
• D- and L- configuration: experimental assignment of optical activity was symbolized with the letter "d" or the "+" sign
(for dextrotatory, right handed or clockwise rotation of the plane of polarized light when viewed toward the light
source) and "l" or "-" (for levorotatory, left or counterclockwise rotation).
• All the amino acids found in proteins are of the L-configuration
• Glycine – only optically inactive among the common amino acids
ACID-BASE PROPERTIES OF AMINO ACIDS
• pKa of the carboxylic group: 2 • pKb of the amino group: 5; pKa = 9
• Under physiologic conditions (about pH 7): amino acids will exist in their zwitterionic forms
• Zwitterion – is a molecule that has a pair of charges, one positive and one negative, such that there is no net charge
↳ Species of amino acid present at pH1 ↳ Known as dipolar ion ↳ Formation depends on a particular condition

Zwitterionic Form of Amino Acids

• Anionic form or least protonated – pH = 9 and ↑ • Cationic form or fully protonated – pH = 2 and ↓
• Diamino Monocarboxylic = (lys|arg|his} • Dicarboxylic Monoamino = (asp|glu)
• Isoelectric pH – take the average value of the pKa before and after the formation of zwitterion
• Isoelectric Point (pI) – pH at which an amino acid is electrically neutral – that is, where the sum of all the positive
charges equals the sum of the negative charges
• For simple amino acid: • For basic amino acid • For acidic amino acid
(𝒑𝑲𝒂𝑪 + 𝒑𝑲𝒂𝑵) (𝟐 𝒉𝒊𝒈𝒉𝒆𝒔𝒕 𝒑𝑲𝒂) (𝟐 𝒍𝒐𝒘𝒆𝒔𝒕 𝒑𝑲𝒂)
𝒑𝑰 = 𝒑𝑰 = 𝒑𝑰 =
𝟐 𝟐 𝟐

• Henderson-Hasselbalch Equation:
[𝑯𝑨]
𝒑𝑯 = 𝒑𝑲𝒂 − 𝒍𝒐𝒈 [𝑨− ]

9
SEPARATION TECHNIQUES
Gel Electrophoresis
– common technique for separating amino acids (or proteins) involved placing a mixture in the center of a gel and
then applying a strong voltage across this gel
– based on the fact that amino acids or proteins that carry a net positive charge at the pH at which the separation
1 is done will move toward the negative electrode, whereas those with a net negative charge will move toward the
positive electrode
– Purification Methods: Isoelectric Focusing (Electrofocusing) and Sodium Dodecyl Sulfate Polyacrylamide Gel
Electrophoresis (SDS-PAGE)
Paper and Thin-Layer Chromatography (TLC)
2 – based on polarity of amino acids
– difference in solubility of amino acid between 2 immiscible solvents
Paper Electrophoresis
3
– acid-base properties of amino acids with the use of an electri field
Ion-Exchange Chromatography
4 – based on coulombic/ionic interactions or affinity to the ion exchanger Cation Exchanger – Polysulfonic Resins
– acid-base properties og amino acid with the use on ion-exchange resin Anion Exchanger – Polyamine Resins
Size Exclusion Chromatography
5
– based on proteins’ Stokes Radius, or the diameter of sphere that they occupy when they tumble in solution
PEPTIDE BONDS
– forms proteins by amide linkages between neighboring amino acids
Dipeptide 2 Amino Acids
Tripeptide 3 Amino Acids
Oligopeptide Few Amino Acids
Polypeptide 10 to 2000 Amino Acids
Dalton Unit of Molecular Weight of Polypeptide
• Condensation Reaction – reaction involved in the formation of peptide bonds
• Amino Terminus – left-most side of peptide (N-terminal)
• Carboxyl Terminus – right-most side of peptide (C-terminal)

STRUCTURAL LEVELS OF PROTEINS – proposed by Kai Linderstrom-Lang

– Sequence of amino acids in a protein

Primary (1o)

– Regular, repeating folding pattern stabilized by hydrogen bonds between amino acids or
peptide groups close together in a sequence
Forms: Super Secondary Structures
Secondary (2o) Right-handed Alpha Helix (helix + beta structure):
Polyproline Type II Helix (collagen) Helix-turn-helix
Parallel Beta-pleated Strand/Sheet Leucine zipper
Antiparallel Beta-pleated Strand/Sheet Zinc finger

10
Segments of secondary structure fold together in 3-dimensions, stabilized Fibrous Proteins
Tertiary (3o)
by interaction often far apart in the sequence Globular Proteins
Quaternary (4o) Association of separate polypeptide chains
DENATURATION
Native State • Natural, biological conformation of a protein
Denaturation • Results in the unfolding and disorganization of a protein’s structure
Consequences of • Loss of activity • Inability to form crystals
Denaturation • Decreased solubility, precipitate out of solutions
• If the conditions of the denaturation process is removed, it is possible that the protein can
Renaturation re-form to its native conformation (except if there is irreversible denaturation caused by
aggregation)

pKa VALUES OF COMMON AMINO ACIDS | (Campbell and Farrel)

AMINO ACID α –COOH (pK1) α –NH3+ (pK2) RH or RH+ (pKR)
Glycine (gly) 2.34 9.60 -
Alanine (ala) 2.34 9.69 -
Valine (val) 2.32 9.62 -
Leucine (leu) 2.36 9.68 -
Isoleucine (ile) 2.36 9.68 -
Serine (ser) 2.21 9.15 -
Threonine (thr) 2.63 10.43 -
Methionine (met) 2.28 9.21 -
Phenylalanine (phe) 1.83 9.13 -
Tryptophan (trp) 2.38 9.39 -
Asparagine (asn) 2.02 8.80 -
Glutamine (gln) 2.17 9.13 -
Proline (pro) 1.99 10.6 -
Aspartic Acid (asp) 2.09 9.82 3.86*
Glutamic Acid (glu) 2.19 9.67 4.25*
Histidine (his) 1.82 9.17 6.0*
Cysteine (cys) 1.71 10.78 8.33*
Tyrosine (tyr) 2.20 9.11 10.07
Lysine (lys) 2.18 8.95 10.53
Arginine (arg) 2.17 9.04 12.48

TITRATION OF AMINO ACIDS

1 Neutral Amino Acids

11
2 Acidic Amino Acids

3 Basic Amino Acids

SEPERATION OF POLYPEPTIDES
A Acid Hydrolysis – destroy trp, loss, ser, thr Glu → Glu + NH3 Asn → Asp + NH3
B Base Hydrolysis – doesn’t destroy trp but destroys cysteine, cystine, ser, thr
C Enzymatic Hydrolysis – divided into two types of Proleolytic Enzymes
Endopeptidases – (cleave internal peptide bonds) Exopeptidase – (cleave external peptide bonds)
Trypsin – cleaves CO functions of lys and arg Carboxypeptidase A – cleave C-terminal if not arg, lys
Chymotrypsin Pepsin – cleaves CO functions of phe, tyr, trp Carboxypeptidase B – cleave C-terminal if it is arg, lys
Thermolysin – cleaves -NH function of val, leu, ile Carboxypeptidase C – non-specific
Elastin – cleave the CO function of neutrophilic aliphatic a.a

CLASSIFICATION ACCORDING TO ACTIVE AND PASSIVE PROTEINS

Active Protein Passive Protein
• Not pure polypeptide of amino acid • Pure amino acid
• Have the ability to bind with other molecules • Doesn’t have the ability to bind with other molecules
CLASSIFICATION ACCORDING TO CHEMICAL DIFFERENCES
1 Simple Proteins – basically composed of amino acids
• Albumin – constitute the most important group of simple proteins
↳ Water soluble; may be precipitated from solutions of high salt concentration. They are coagulated by heat
• Globulin – insoluble in salt-free water; soluble in dilute salt solution but when concentration to 30-50% saturation
they are insoluble

12
• Histones – hydrolysis products of nucleoproteins and are soluble in acid solution, they usually form complex with
nucleic acid. Its small molecular weight proteins are basic
• Albuminoids – insoluble in water and all neutral solvents and are not coagulated by heat
2 Conjugated Proteins – simple proteins linked to non-protein groups
↳ Hydrolysis products are amino acids + non-protein groups
• Chromoproteins – Pigments (Hemoglobin) • Mucoprotein – Carbohydrate (Mucin)
• Glycoprotein – Carbohydrate (Globulin) • Nucleoprotein – Nucleic Acid (Virus)
• Lecithoprotein – Phospholipids (Lipoprotein) • Flavoprotein – Flavin Nucleotide (Amino A. Oxidase)
• Metalloprotein – Metal (Ferritin) • Phosphoprotein – Phosphate (Casein)
3 Derived Proteins – products of hydrolysis (partial to complete) and conjugated proteins
↳ Coagulated proteins that are formed by heating or treatment of alcohol, acids, bases
↳ Hydrolysis products and modification products of other classes of proteins
Primary Derivative Proteins Secondary Derived Proteins
• No change in molecular weight after heating • Products of more extensive hydrolysis
CLASSIFICATION ACCORDING TO SHAPE
FIBROUS GLOBULAR
• Structural proteins of animals and include the proteins of • Arranged in tight, compact, spherical shape
hair, skin, nails, and connective tissues
• Arranged in parallel along a single axis • Soluble in water
• Insoluble in water and resistant to denaturation • They are proteins of meat, milk, and eggs
• Example: collagen, keratin, myosin, fibrin • More sensitive to denaturation than fibrous proteins
ACCORDING TO BIOLOGICAL IMPORTANCE
1 Body’s main destiny source of nitrogen and sulfur 2 Catalytic and structural functions
3 They make up contractile system of muscles 4 Antibodies – they are the defense system of the body
5 Hormones – regulate the body’s glandular activity 6 Some antibodies that are secretions of bacteria and fungi
7 In the blood, they maintain the fluid balance that are part of the clotting process and transport O2 and lipids
Can act as poisons, such as the venoms in animal bites and stings or toxins, such as the bacterial toxin that produces
8
botulism in humans from eating improperly processed foods
THE DYNAMIC STATE OF PROTEINS

During digestion, proteins in the food are broken down into amino acids which are then absorbed and used to
synthesize the proteins needed in the body – the main source of nitrogen in the diet
CLASSIFICATION OF PROTEINS BY BIOLOGICAL FUNCTION
1 Structural Proteins
• Keratin – primary constituent of skin, hair, nails • Collagen – makes up tendons and developin bones
2 Contractile Proteins – structural components of muscles – for control and relaxation
• Actin – moving filamints in myofibril • Myosin – stationary filaments in myofibril
3 Enzymes – biggest class of proteins – meatbolic proteins – biological catalyst
• Trypsin – hydrolyzes peptides • Amylase – hydrolyzes polysaccharides
4 Hormones – for regulation of metabolism – not all hormones are protein in nature
• Insulin – lowers the effect of free glucose • Growth Hormone – stimulates growth of bones
5 Transport Proteins
• Hemoglobin – transports oxygen in blood of vertebrates • Iron-binding Globulin – transports iron in the blood

13
• Serum Albumin – transports fatty acids in the blood • Myoglobulin – transports oxygen in muscle cells
6 Protective/Defensive Proteins – defensive mechanism
• Antibodies – prevent invading viruses and bacteria – forms complexes with foreign proteins
• Fibrinogen – precursor of fibrin in blood clotting • Thrombin -- component in the clotting mechanism
7 Storage Proteins – stores energy
• Casein – milk protein • Ovalbumin – egg white protein • Zein – corn seed protein • Gliadin – wheat seed pr.
8 Toxins
• Snake Venom – enzymes that hydrolyzes phosphoglycerides • Ricin – toxic protein of cottonseed
• Clostridium Botulinum Toxin – causes bacterial food poisoning
DNA A – Chain B – Chain
Human Thr–Ser–Ile –Thr
Bovine Ala–Ser–Val –Ala
Hog Thr–Ser–Ile –Ala
Sheep Ala–Gly–Val –Ala

STRUCTURES OF PROTEINS

Covalent Forces/Interaction – strong bonds like peptide bonds, amide bonds, ester bonds, disulfide bonds (formed
1 between 2 adjacent cysteine residues; strong covalent bond that confers considerable rigidity on the protein
molecule
Noncovalent Interaction
• H-Bonding -- weaker bond than ionic bond; electrostatic attraction between electropositive hydrogen atom from
one molecule and an electronegative nitrogen, oxygen or fluorine from a neighboring molecule
H – donors H – acceptors
2 >NH –S–S–
–OH C=O
–NH2 –COO–
+
–NH 3
• Electrostatic Forces/Ionic Interaction/Salt Bond – moderately strong bond – bonding between opposite charge

14
Positive (+) Charge Negative (–) Charge
α – amino α – COOH
ε – amino (lys) β – COOH (asp)
Guanidinium (arg) γ – COOH (glu)
• Hydrophobic Interaction – these bonds arise from a tendency of certain groups to exclude water molecules from
2 their vicinity. Weaker bonds produced by interaction of nonpolar sidechains and caused by mutual repulsion of the
solvent
• Van der Waal’s Forces -- produced by interactions between polar sidechains
Attractive Forces Repulsive Forces
• Dependent on the distance between residues • underlying forces in steric hindrance and bulky groups
• Can exist between ser, aromatic, nonpolar residues in proteins

LEVELS OF PROTEIN STRUCTURE

I Primary Structure of Proteins – refers to the identity, number, sequence of amino acids in the polypeptide chain
↳ Polymer of amino acids ↳ Stabilized peptide bonds – covalent bonds
• The amino acid sequence for any given protein is always identical and the slightest variation in this sequence may
have dramatic biological consequences
II Secondary Structure of Proteins – is the shape in which the long amino acid chain exits
↳ The folding of parts of polypeptide chains into specific structures held together by hydrogen bonds
↳ Noncovalent interactions within (1 polypeptide will form H-bond with itself) or between (2 or more) the polypeptide
backbone. It is fibrous in nature, insoluble. Without H-bonds, there is no secondary structure
Essential Features of some Secondary Structures
A Alpha–Helix
• Backbone is arranged like a coil in helical conformation • Coiling occurs within a single polypeptide chain
• Only intrapeptide H-bonding between –NH and C=O • 3.6 amino acids per turn (5.4 A) – repeat distance
• Rise per reside is 1.5 A • 13 atoms involved in one turn (3.6 amino acids)
• Each H-bond is weak, but the large number gives it stability • May be right (clockwise) or left (counter) handed
• The amino acid sidechains can be accommodated as they stick out into space away from the polypeptide chain
• Every amide hydrogen and carbonyl oxygen associated with the peptide backbone is involved in a hydrogen bond
when the chain coils into an α-helix. These H-bonds lock the α-helix into place
• Every carbonyl oxygen is hydrogen bonded to an amide hydrogen four amino acids away in the chain
• The hydrogen bonds of the α-helix are parallel to the long axis of the helix
• The polypeptide chain in an α-helix is right-handed. It is oriented like a normal screw
• Clockwise – goes into the wall • Counterclockwise – comes out of the wall
Variation of the Alpha-Helix:
• Presence of proline • Presence of adjacent similarly charged group • Presence of adjacent bulky group
• Proline as Helix-Breaker – the amino group is part of the ring and is unable to fit into the rigid geometry required by
the alpha-helix when incorporated into a peptide. It lacks the alpha-amino proton for H-bond formation

15
B Beta-pleated Sheet
• Two or more polypeptide chains • H-bonding is interpeptide • Less bulky amino acids present
• R-groups or sidechains of amino acid residues live above or below the zigzag planes
• Bonds occur between two or more polypeptide chains which give an overall pleated shape to the structure
• Polypeptide chains are nearly completely extended, and hydrogen bonds between C-O and N-H groups are at right
angles to the long axis of the polypeptide chains

• Secondary protein structure in which the backbone of two protein chains in the same or different molecule is held
together by hydrogen bonds
• The orderly alignment of protein chains is maintained by intermolecular or intramolecular hydrogen bonds
Two Types of Beta-Pleated Sheet:
1 Parallel Beta-Pleated Sheet
• When polypeptide chains are in the same direction and start with the same terminals
• They start with different terminals and end with different terminals
2 Antiparallel Beta-Pleated Sheet
• When polypeptide chains are run in opposite direction and starts and ends with different terminals
• Ex: Silk Fibroin – contains sections that consists of only alanine (25%) and glycine (42%)
C Three-Stranded Helix
• Found in collagen • Has no alpha helix due to high glycine and proline content
D Random Coil
• Neighboring R-groups repel each other
• There is no consistent relationship between the amino acids and their stabilizing bonds
Preferred Secondary Structures of Some Amino Acids
1 Helix Forming – ala, trp, tyr, met, val, gln, leu, phe, cys, his, asn
2 Non-Helix Forming (Destabilizing Alpha Helix) – ser, thr, asp, gly, ile, glu, lys, arg
3 Beta-Pleated Structure (Due to small sidechain) – gly
4 Random Coil – glu, asp, lys, arg
5 Alpha-Helix Breaker – pro
6 Cross-Linkages (Intramolecular) – cysteine
III Tertiary Structure of Proteins
• Tendency for a polypeptide chain to undergo extensive coiling or folding to produce a complex rigid structure
• If the helical coil is intertwined into roughly a sphere, the protein is a globular protein, whereas the winding together
of helical coils forms long strands, it give rise to fibrous proteins
• Shape is maintained by salt bonds (electrostatic attraction), hydrogen bonds, disulfide bonds, van der waal’s forces
and hydrophobic interactions (major force in maintaining the shape or conformation of proteins)
• Hydrophobic interaction allows a conformation such that hydrophobic R-groups are on the interior and maximally
shielded from water and as such lower the free energy content of the protein
• Breaking one or more bonds causes denaturation • 1 Polypeptide = 1 Tertiary
Globular Rod-like
Ribonuclease Fibrinogen
Ovalbumin Collagen
Serum Albumin Myosin

16
IV Quaternary Structure of Proteins
• Spatial relationship and interactions between subunits in a protein that has more than one polypeptide chain
• The association of similar or dissimilar protein units into oligomers (small polymers). The Sub-units are held together
by non-covalent forces. The same attractive forces that stabilize the tertiary structure maintains quaternary structure

Classification of Proteins in Quaternary Level

• 2 Sub-units – Dimer • 3 Sub-units – Trimer • 4 Sub-units – Tetramer
• Homogeneous – if sub-units are identical • Heterogeneous – if sub-units are not identical
Examples No. of Protein Sub-units
Insulin 2
Thrombin 3
Hemoglobin 4
Arginine Decarboxylase 5
Glucose-6-Phosphate Dehydrogenase 6
Lipoic Reductase Transacetylase 60
Poliomyelitis Virus 130
V Quintenary Structure
• Groups of proteins (in quaternary level), each with distinctive biological activity existing together as protein-protein
aggregates. Only proteins exist in this level. Ex. Pyruvate Dehydrogenase

DENATURATION OF PROTEINS
• The loss of the secondary, tertiary, and quaternary structures of a protein by a chemical or physical agent that leaves
the primary structure intact
• Conversion of a protein to an unfolded or altered state
• Alteration from the naturally ordered conformation to a randomly structure molecule
• Disrupts or breaks the attractive forces maintaining the native configuration
• Native State – highly ordered conformation in which the biological activity of the protein is manifested
• As heat is applied, the H-bonds maintaining the secondary structure are disrupted, and the protein structure becomes
disorganized. Thus, the protein is denatured.
Results of Denaturation
1 Coagulation
• When chains unfold and become entangled • Proteins are no longer soluble • Usually no reagent is added
2 Precipitation
• Becomes insoluble upon addiction of reagent
Types of Denaturation
Reversible Irreversible
• Denaturation is mild • Denaturation is drastic
• Can be returned to original nature • Can no longer go back to its original form
• Ex: Ironing Hair • Ex: Cooking Egg

17
Consequences of Denaturation
1 Loss of biological activity – destroys functions
2 Loss of solubility – becomes insoluble
3 Loss of crystallizability – becomes soluble
4 Cleavage of H-bonds – breaks
5 Increased number of ionizable groups
6 Increase in susceptibility to hydrolysis by proteolytic enzymes
Denaturing Agents for Proteins
Denaturing Agents How the Reagent May Operate
Disrupts hydrophobic interactions and H-bonds by making molecules vibrate violently
Heat
which produces coagulation
Causes the violent vibrations of molecules that disrupt H-bonds and hydrophobic
Microwave Radiation
interactions
Ultraviolet Radiation Probably operates much like the action of heat
Violent Whipping/Shaking Causes molecules in globular shapes to extend to longer lengths and then entangles
Soaps/Detergent Disrupts hydrophobic interaction
Organic Solvents (ethanol, May interfere with H-bonds because these solvents can also form H-bonds or can
acetone, 2-propanol) disrupt hydrophobic interactions. Quickly denature proteins in bacteria, killing them.
Disrupt H-bonds and salt bridges. Prolonged action leads to actual hydrolysis of peptide
Strong Acids & Bases
bonds
Salts of Heavy Metals Cations combine with SH-groups (Sulfhydryl) and form precipitates. These salts are all
2+, + 2+
(salts of Hg Ag , Pb ) poisonous. It Destroys salt bridges and disulfide bonds

CHEMICAL PROPERTIES OF PROTEINS

1 Coagulation and Precipitation – proteins are coagulated by (A) – (F)
A Heat -- egg albumin coagulates upon heating
Mineral Acids (like HCl and HNO3) – Casein in milk is precipitated by addition of HCl
↳ This is the principle involved in Heller’s ring test for coagulated protein in urine. If urine is slowly poured down
B
on the sides of a tube containing HNO3 and a white precipitate is formed at the interface of the two liquids, the
presence of coagulated protein is indicated
Salts of Heavy Metals, like Mercuric Chloride, Silver Nitrate, forming the insoluble metallic proteinates, the
C deficiency of disinfectants depends on this property of coagulating the protein of the organism destroying its life.
The use of egg whites as an antidote for metallic poisoning is based on the ability of the egg whites to be coagulated
by the metallic ion thus taking it out of the field of action.
D Neutral Salts – Ammonium Sulfate & Magnesium Sulfate – these substances are used to salt out proteins
E Alcohol – destroys cell wall and owns its disinfecting power to this property in the form of 50-70% sol’n
Alkaloidal Reagents such as Picric Acid, Tannic Acid, Phosphotungstic Acid, Phosphomolybdic Acid
F ↳ Tannic Acid is used to relieve diarrhea, to check secretions in the treatment of burns because it produces and
astringent effect on the tissues and prevents absorption of toxins.
↳ Phosphotungstic Acid is used to precipitate and remove proteins in blood analysis
2 Hydrolysis – proteins are hydrolyzed by acids, alkalines, enzymes
Color Reaction of Proteins -- proteins give various color reactions which is not specific for protein molecules, but
3
for certain groups of the component amino acid
Biuret Test – alkaline solution of protein treated with CuSO4 solution results in the production of a rose-ink to
violet coloration. This is due to the presence of peptide linkage (-CONH). All substances therefore containing this
A linkage respond positively to this test. The smaller peptides and amino acids show no coloration. This test serves
as a good index for determining the extent of protein hydrolysis. Their hydrolytic products give colors ranging from
violet to pink depending upon the extent to which hydrolysis has progressed.
Xanthoproteic Reaction – proteins when treated with HNO3 turn yellow and change to orange when neutralized
B
with an alkali (NH4OH). This is due to the nitration of the benzene ring (-C6H5) present in the structure of such amino

18
acids such as tyrosine, tryptophan, phenylalanine (the yellow stain on the skin produced by HNO3 is due to this
B
reaction). Nitration of benzene ring results to yellow orange when salt
Millon’s Test – is made by dissolving mercury in HNO3 – hence it consists of mercurous and mercuric nitrates.
When added to a protein solution, the protein is precipitated as mercury salt. On heating, the precipitate turns
C
flesh to red color if proteins containing tyrosine is present . This reaction is due to the phenol group contained in
tyrosine. Any substance therefore containing this group will give a positive result
Heller’s Ring Test – if nitric acid is introduced into a test tube as a lower layer beneath a solution of protein, a white
D precipitate of coagulated protein appears at the junction of the two layers. This test is commonly used for detecting
albumin in urine
Ninhydrin Reaction – alpha amino acids given with triketohydrindene hydrate (Ninhydrin) results in a blue to
E
violet-red color. This is a very delicate test for both the proteins and amino acids
Sulfur Test – solutions of protein containing cystine, cysteine or methionine, when heated with NaOH, splits up the
F sulfur to form Na2S. When treated with lead acetate it produces a black precipitate due to the formation of Lead
(II) Sulfide
Hopkins-Cole Reaction – when protein mixed with glyoxylic acid (CHOCOOH) is treated with sulfuric acid, a violet
ring is produced at the point of contact of the two solutions. This is due to the presence of an indole nucleus in the
G
tryptophan component. The tryptophan condenses with the aldehyde to form the colored compound
Glyoxylic Acid – made by the reduction of oxalic acid with magnesium powder
Nitroprusside Test -- proteins containing cysteine give a red color with sodium nitroprusside in dilute ammoniacal
H
solution
Molisch’s Test – (for sugar) if a protein solution is treated with Molisch’s reagent and layered with sulfuric acid, a
I
violet ring will develop at the junction of the liquid layers if cycloprotein is present
J Liebermann’s Test – protein + HCl turns violet due to the presence of tryptophan
Sakaguchi’s Test – in alkaline solution, protein containing arginine gives a red colored Alpha-Naphthol and Sodium
K
Hypochlorite. This is a test for arginine
Sullivan’s Test -- test for cysteine and cystine after reduction. It gives a red color with Sodium 1,2-naphtho-quinone-
L
4-sulfonate
Pauly’s Reaction – histidine and tyrosine in alkaline solution when made to react with diazotized sulfanilic acid
M
gives a red color

COMPLETE PROTEINS IN THE DIET

• The composition of dietary protein must match the requirements for the formation of body protein, if one or more
necessary amino acids are not available at the time of synthesis of vital protein, then the protein is not made. Protein
synthesis is an “all or nothing” event
• Dietary proteins are rated in terms of biological value on a percentage scale. Complete Proteins have a high biological
value and supply all the required amino acids in the amounts required for normal growth
BIOLOGICAL VALUE OF DIETARY PROTEINS
FOOD BIOLOGICAL VALUE (%)
Whole Hen’s Egg 94
Whole Cow’s Milk 84
Fish 83
Beef 73
Soybeans 73
White Potato 67
Whole Grain Wheat 65
Whole Grain Corn 59
Dry Beans 58
• Note that hen’s egg, cow’s milk, and fish provide high biological value proteins. Plant proteins vary more in biological
value than animal proteins. However, not all plant proteins are deficient in the same amino acids. Gliadin, a wheat
protein, is low in lysine; Zein, a corn protein is low in both lysine and tryptophan. Societies that large amounts of corn
and wheat products must have other sources of lysine

19
• Vegetarians must carefully choose their food so that all of the essential amino acids are available. For example, beans
are high in lysine, whereas wheat is low in lysine. Similarly, wheat is high in cysteine and methionine, whereas beans
are low in these two. By eating both, the vegetarian increases the percentage of the usable proteins.
EFFECTS OF PROTEIN DEFICIENCY
1 Retardation of growth in children and loss of weight in adults
2 Hemoglobin is diminished to anemia
3 Production of fatty liver due to lack of lipotropic amino acid and methionine
4 Plasma proteins are decreased leading to edema
5 Resistance to infection becomes low due to lack of antibodies (gamma-globulin)
Endocrine disturbances like amenorrhea (failure to menstruate) due to deficient gonadotropin, a protein hormone
6
from the anterior pituitary
7 Enzyme production becomes less, leading to impairment of functions, especially the gastrointestinal tract

ENZYMES
• Are secreted by living cells and are complex organic chemical compounds with definite structure. Like the inorganic
catalysts, they have the remarkable property of speeding up chemical reactions without being themselves affected in
the process. As a result, they can be used over & over again. Also, the largest & most highly specialized class of proteins
• Metabolism takes place very rapidly in living organisms with many individual reactions complete in milliseconds or
less. This fast rate of metabolism is only possible because of the enormous increase in the rate of chemical reactions
brought about by enzymes. Without these enzymes, the chemical reactions would not take place at a fast-enough rate
to maintain life. Thus, the absence of even one of the enzymes in the body can have serious consequences.
• Are water-soluble globular proteins that can vary from a molecular weight of 12,000 to 1 million. The enzyme
molecule may consist of a fairly simple single polypeptide chain or it maybe a more complex molecule composed of
several polypeptide chain of 124 amino acids
CHARACTERISTICS
• Endowed with catalytic power and specificity. Enzymes are highly specific
Carbonic Anhydrase
1 CO2 +H2O --------------------> H2CO3
↳ In the presence of Carbonic Anhydrase, formation is increased 106 times
↳ The above reaction can only be catalyzed by Carbonic Anhydrase
2 • Activity is regulated 3 • Involved in transformation of energy
• Some enzymes are simple proteins and others are conjugated (contain nonprotein past) that require a nonprotein
group for their biological activity
• Simple Proteins – entirely amino acids in composition • Nonprotein Part – prosthetic part
Examples of enzymes that are composed solely of amino acids:
• Pepsin • Trypsin • Chymotrypsin • Ribonuclease • Urease • Most Hydrolases
SOME IMPORTANT DEFINITIONS
Apoenzyme – the term used to describe the protein part of the enzyme molecule
1
↳ Catalytically inactive ↳ Require cofactors to become catalytically active ↳ Pure protein
Cofactors – nonprotein component required by some enzymes to be able to perform their catalytic functions
2 ↳ Maybe metal ion (activators) or an organic molecule called coenzymes
↳ Some enzymes require both types of cofactors ↳ Inorganic temporary
3 Activators – when the cofactor of an enzyme is a metal ion Some Activators: Mg+2,Zn+2,Fe+3,Mn+2
Coenzymes – small nonprotein organic molecule which aid enzymes in performing their catalytic functions
4
↳ Organic Temporary Ex: NAD, FAD, CoQ, PP, TPP
5 Holoenzyme – catalytically active enzyme – cofactor complex ↳ Nonprotein + Apoenzyme = Holoenzyme
6 Metalloenzyme – enzymes requiring metal ions
7 Antienzyme – inhibits the action of an enzyme substrate
8 Prosthetic Group – cofactor that is tightly bound to the protein ↳ Organic/Inorganic Permanent
9 Isoenzyme – occurs in multiple forms within an organism
Proenzyme (Zymogen) – name given to the inactive form of an enzyme
10
↳ must be activated to be able to perform their catalytic functions

20
• Pepsinogen – product of the gastric mucosa – Is converted to active pepsin by HCl present in the gastric juice
10 • Trypsinogen – product of the pancreas – changed into active trypsin by enterokinase in the duodenum
• Chymotrypsinogen – also produced by the pancreas – when this proenzyme comes in contact with trypsin in the
small intestine, it is converted into active chymotrypsin
11 Substrate – the chemical substance/s upon which the enzyme acts ↳ Undergoes change
12 Active Site – the specific are of the enzyme to which the substrate attaches during the reaction
NOMECLATURE OF ENZYMES
1 Some enzymes have been chemically uninformative names. Ex: pepsin, rennin, trypsin, chymotrypsin
Some enzymes have been named by adding the suffix –ase to the
2 • Arginase – catalyze the hydrolysis of arginine to ornithine and urea • Lipase – lipids
• Maltase – catalyze the hydrolysis of maltose to glucose • Hydrolase – catalyzes hydrolytic rxn.
• Dehydrogenase – catalyzes the removal of H from substance • Decarboxylase – removal or carboxyl groups
Some names are combinations of several designations • Malate Dehydrogenase
3
• Xanthine Oxidase – an enzyme which catalyzes oxidation of xanthine • Succinate Dehydrogenase
Both the names of the substrate and of the reaction type • Lactate Dehydrogenase • Pyruvate Decarboxylase

ENZYME CLASSIFICATION
I According to the classification by the Enzyme Commission of I.U.B (International Union of Biochemistry)
6 Major Classes
Code Class Name Reaction
1 Oxido-Reductases Redox reactions
2 Transferases Transfer of functional groups except H
3 Hydrolases Hydrolytic reactions
4 Lysases Catalyze the non-hydrolytic removal of groups usually from double bonds
5 Isomerases Interconversion of optical, geometrical functional, or stereoisomers
Catalyzes the joining of 2 molecules with the breaking of a pyrophosphate bond or
6 Ligases
catalyze formation of bonds with ATP cleavage
II According to its presence in the cell
1 Constitutive Enzymes – part and parcel of the cell 2 Inductive Enzymes – induced to be present in the cell
III According to Substrate Specificity
Absolute Specificity – enzyme catalyzes on specific Relative Specificity – enzyme acts on a particular kind
substrate. of covalent bond in closely related substrate
1 2
Ex: fumarase – act on fumaric acid only Ex: proteolytic enzymes – hydrolyze peptide bonds of
Ex: sucrose – acts on sucrose only polypeptides
MODE OF ENZYME ACTION – HOW ENZYMES WORK?
• General Statements of Enzymatic Reaction: E+S → ES → E+P E= enzyme, S= substrate, ES = enz-sub complex
• Only a portion of the enzyme molecule is concerned with its catalytic activity. This region is called active/catalytic site
• Active/Catalytic Site – the site directly involved in catalysis • Small portion of the enzyme which is hydophobic
• A-B-C Contact Residues – have contact with substrate

↳ Amino acids or proteins which orient properly the

substrate at the active zone
↳ Plays an active site by donating or withdrawing electron
from the functional groups of the substrare
• X-Y-Z Auxillary Residues – don’t have contact with
substrate
↳ Helps maintain and stabilize the functional conformation
of the active site

21
2 THEORIES OF ENZYME ACTION
1 Lock and Key Theory (Rigid Model by Fischer)
↳ The active site is often in a fold or cleft in the enzyme into which the substrate molecules fit
↳ This let Fischer to suggest the lock-key hypothesis in which close binding of the substrate to the active site
↳ The active site is rigid – where the shape of the enzyme active site is designed for specific substrate
2 Induced-Fit Theory (Flexible model by Koshland)
↳ Hand and glove; where the active site is flexible and changes shape to accommodate the substrate as it approaches
↳ The substrate induces the enzyme to undergo some folding, unfolding or rotation about peptide bonds so that contact
amino acids are correct spatial orientation.

ENZYME INHIBITION / INHIBITION OF ENZYME ACTIVITY

• Normal Reaction *Activation – process that initiates or
increase the action of enzyme
• Inhibit – to restrain, prohibit, hold, suppress. Makes an
active enzyme less active or inactive
• Inhibitors – substances that specifically decrease the
rate of enzymatic reactions; making the enzyme less active
• Specific Inhibitors – affect one enzyme or group. In this
group are most poisonous substances, such as Cyanide
which inhibits the activity of Cytochrome Oxidase enzyme
• Non-specific Inhibitors – affect enzymes in the same manner
Heat Changes in pH Strong Acid Alcohol Alkaloidal Reagents = all can denature proteins
• Activators – are inorganic substances that tend to increase the activity of an enzyme
• Mg2+ – inorganic activator of the enzyme phosphatase • Zn2+ – activator for the enzyme carbonic anhydrase
Examples of Inhibitors
1 Toxic Gases – used in biological weapons and inhibit key enzymes
2 Drugs (antibiotics/antimetabolites) – inhibit a variety of enzymes necessary for bacterial growth
Organophosphorus Compounds – toxic ingredients of insecticides
3 ↳ Potent irreversible inhibitors of enzymes that have active seryl residues at their catalytic site
↳ Inhibitors of acetylcholinesterase – key enzyme involved in the transmission of the nerve impulse at the
neuromuscular junction and parts of the central nervous system
4 Heavy Metal Ions (Pb2+,Hg2+,Ag2+) – inhibit the activity of some enzymes by binding to thiol/sulfahydryl grp. (-SH)
5 Carbon Monoxide – forms stable complex with hemoglobin
6 Cyanide (CN-) – kills by killing cytochrome oxidase – present in all aerobic cells, for intracellular respiration
Advantages: • Means of Metabolic Control • Enzyme Activity is Regulated
Disadvantages: • Enzymes are destroyed; no more catalysis or catalytic activity
Types of Inhibition
• Irreversible – Inhibitors become covalently bonded/linked to the enzyme
E+I → EI ES+I → ESI ESI and EI complexes cannot break down to regenerate E
↳ Enzyme activity decreases with time and eventually reaches zero
Examples of Irreversible Inhibitors CN- CO Organophosphorus Compounds
• Reversible – form EI or EIS or both but so reversibly allowing E to be functional again
E+I → EI ES+I → ESI Allows E to be functional
3 Types of Reversible Inhibition
• Competitive – the S and I compete for the same active site E+I → EI competes with E+S → ES
1 ↳ Binds to the active site of the enzyme surface thus preventing the binding of substrate
↳ Inhibition can be relieved by increasing the concentration of S
↳ S and I – compete for the same site because they have similar chemical structures

22
p-aminobenzoic acid Sulfanilamide
(bacterial growth factor) (antibacterial factor)
1

↳ The sulfa drug; sulfanilamide prevents bacterial growth by competitive inhibition

• Non-competitive – binds to some other portion of the enzyme surface
2 ↳ May sufficiently alter the tertiary structure of the enzyme so that its catalytic effectiveness is slowed down
↳ I – does not bind to the site but reacts to another site causing distortion or deformity of the E, thus the ES is not
formed at a normal rate and once forms the ES complex, it does not decompose at the normal rate to yield P
• Uncompetitive – I does not combine with free E but combine reversibly with ES complex to form inactive ESI
3
↳ ESI – cannot undergo further reaction to yield the normal products
Specificity of the Enzyme-Substrate Complex
• Enzyme Specificity – ability of an enzyme to bind only one, or very few substrates, thus, catalyzing only a single rxn
• Absolute Specificity – catalyzes the reaction of only one substrate; lock and key Ex. Urease, Sucrase
• Grouped Specificity – catalyzes processes involving similar molecules containing the same functional group; induced-
fit. Monosaccharides to Monosaccharide, not polysaccharide. Ex. Hexokinase – catalyzes phosphate
• Linkage Specificity – catalyzes the formation or breakage of only certain bonds in a molecule; induced-fit
Ex. Pepsin, Amylase – breaks the alpha glycosylic (starch, sucrose, polysaccharide) but not beta glycosylic (cellulose) –
we can’t digest grass directly because it is made up of cellulose
• Stereochemical Specificity – enzyme that can distinguish one enantiomer form the other same; induced fit (D-, L-)

COENZYMES
Small nonprotein organic molecule which aid enzymes in performing their catalytic activity
Binds reversibly to the enzyme and its functions as a second substrate for the enzyme
Derived from B-vitamins (H2O soluble vitamins) Coenzyme precursors are vitamins
Derived from Vitamin B complex Temporarily attached
Vitamins – small organic molecules that are essential to maintain health but that either cannot be synthesized by the
body or are synthesized in insufficient amounts
Must be present in the diet for proper cellular function, growth and reproduction
When vitamins are lacking the diet, specific deficiency diseases result

CLASSIFICATIONS OF ENZYMES
— +
I Coenzyme for transfer of e or H
COENZYME VITAMIN SOURCE
NAD+/NADP Niacin (B5)
FAD/FMN Roboflavin (B2)
Lipoic Acid B-complex
CoQ
II Coenzymes for Group Transfer
COENZYME VITAMIN SOURCE GROUP
TPP Thiamine (B1) Acyl, Aldehyde
Lipoic Acid B-complex Acyl
CoA/CoASH Pantothenic Acid (B3) Acyl, Acetyl
PP Pyridoxine (B6) Amino
Biocytin Biotin (B7) CO2
Methyl, Methylene, Formyl,
THF Coenzyme Folic Acid (B9)
Formimino (-CH=NH)
23
Cobamide Enzyme Cobalamin (B12) Alkyl
III Coenzymes used as Energy Rich Source
OXIDIZED FORM REDUCED FORM
NAD+ NADH
FAD FADH2
CoQ CoQH2

THE SEQUENCE SPECIFICITIES OF SOME PROLEOLYTIC ENZYMES

CARBOHYDRATES

24
CLASSIFICATION OF 20 COMMON AMINO ACIDS
Aliphatic Amino Acids
Glycine|(Gly)| G Alanine|(Ala)| A Proline|(Pro)| P

Aliphatic Amino Acids (Branched)

Valine|(Val)| V Isoleucine|(IIe)| I Leucine|(Leu)| L

Aromatic Amino Acids

Phenylalanine|(Phe)| F Tyrosine|(Tyr)| Y Tryptophan|(Trp)| W

Basic Amino Acids

Lysine|(Lys)| K Arginine|(Arg)| R Histidine|(His)| H

Acidic Amino Acids Amide-Containing Amino Acids

Aspartic-Acid|(Asp)| D Glutamic-Acid|(Glu)| E Asparagine|(Asn)| N Glutamine|(Gln)| Q

Sulfur-Containing Amino Acids Hydroxyl-Containing Amino Acids

Cysteine|(Cys)| C Methionine|(Met)| M Serine|(Ser)| S Threonine|(Thr)| T

25
CLASSIFICATION OF PROTEINS (Based on Physicochemical Properties)
A Simple Proteins – naturally occurring proteins that yield only alpha amino acids or their derivatives on hydrolysis
Type Example of Occurrence
Albumins • Ovalbumin in egg • Serum albumin in blood
Globulins • Serum Globulin
Glutelins • Glutenin in wheat
Prolamines • Zein in corn • Gliadin in wheat
Albuminoids • Keratin in Hair & Horny Tissue • Elastin in Tendons & Arteries • Collagen in Skin and Tendons
Histones • Histones and Hemoglobin
Protamines • Salmin and Sturin in fish sperm
B Conjugated Proteins – Contains Amino Acids and Prosthetics (additional inorganic or organic groups)
Type Example of Occurrence
Nucleoprotein Protein + Nucleic Acid – plays role in Genetic Information
Lipoprotein Protein + Lipid (Example: Fatty Acids, Lecithin, Cephalin To Form VLDL, LDL, HDL)
Glycoprotein & Protein + Carbohydrate (Example: Mucin, Immunoglobulin, Complements, Enzymes)
Proteoglycans
Chromoprotein Protein + Pigments (Example: Hemoglobin, Cytochromes, Rhodopsin)
Metalloprotein Protein + Metal (Example: Fe in Ferretin, Cu in Ceruloplasmin, Zn in Carb Anhydrase)
Phosphoprotein Protein + Phosphoric Acid (Example: Casein in Milk, Ovovitellin in Egg Yolk)
C Derived Proteins – from simple/compound proteins by denaturation or by chemical reaction such as hydrolysis
Type Example of Occurrence
Primary Substances formed during hydrolysis of proteins, but with slight alteration
Insoluble products of the action off water, very dilute acids or Enzymes (Myosin from Myosin,
Proteans
Edestan from Elastin, Fibrin from Fibrinogen)
Soluble products in very weak acids and alkalis, but not soluble in neutral solvents (acid and alkali
Metaproteins
metaproteins)
Coagulated Insoluble products resulting from the application of heat to their solutions or to the action of alcohols
Proteins on proteins (cooked egg albumin, cooked meat)
Secondary Substances formed during the progressive hydrolysis of proteins
Proteoses Highest molecular weight group
Peptones Lower molecular weight than proteoses
Peptides Small hydrolytic fragments containing 2-20 Amino Acids or so – lightest
*Complete Hydrolysis: Protein → Protean → Metaprotein → Proteose → Peptone → Peptide → Amino Acid
Based on Molecular Length and Shape
Type Example of Occurrence
Thread like fibrils or long fibers – insoluble in water – structural roles
Fibrous Axial ratio is less than 10
Example: Collagen, Elastin, Keratin
Tightly folded into spherical or ovoid shapes – soluble in water – metabolic roles
Globular Axial ratio is greater than 10
Example: Albumin, Globulin, Enzymes, Hemoglobin

TESTS FOR PROTEINS AND AMINO GROUPS

Test Reactive Group Positive Visible Result
Ninhydrin Amino Acids and Free Amino Group Deep-Violet Sol’n (Ruhemman’s Purple)
Biuret Peptide Linkages Rose Pink/ Violet/ Purple Sol’n
Ehrlich & Hopkin’s Cole Tryptophan Reddish-brown to Violet ring at the junction of two
(Glyoxylic Acid) liquids
Sakaguchi Arginine Red/ Orange Sol’n

26
Xanthoproteic Tyrosine, Tryptophan Yellow/ Orange Sol’n
Pauli Diazo Tyrosine, Histidine Red = Tyrosine Yellow = Histidine
(Pauly Reaction)
Millon-Nasse Tyrosine White ppt. turning into deep red or old rose ppt.
Bromine Water Tryptophan Pinkish-lavender or Violet in the alcohol layer
Basic Lead Cystein, Cystine (sulfur-containing) Black ppt.

EXAMPLES OF PROTEINS AND RELATED DISEASES

Example Description Disease
Most abundant protein in the body – long, rigid Ehler-Danlos Syndrome–inheritable defects in
structure in which three polypeptides are wound the metabolism of fibrillary collagen molecules
Collagen
around one another in a rope-like triple helix – for Osteogenesis Imperfecta – Brittle Bone
structural purposes Syndrome
Connective tissue protein with rubber-like A1-Antitrypsin Deficiency – leads to
Elastin properties Emphysema and cirrhosis due to the inability of
inhibiting elastase by enough antitrypsin
Hemeprotein present in heart and skeletal Sickle Cell Disease (Hemoglobin S Disease) –
muscle, functions both as a reservoir for oxygen, genetic disorder caused by a single nucleotide
and as a carrier for oxygen alteration in B-globin chain
Hemoglobin (heme = protoporphyrin IX and ferrous ion) Hemoglobin C Disease
Hemoglobin SC Disease
Methemoglobinemas
Thalassemias
Found exclusively in red blood cells, where its
Myoglobin main function is to transport oxygen from the -
lungs to the capillaries of the tissues
Peptide hormone made by the beta cells of the
Insulin pancreas that allows body to use sugar from -
carbohydrates
Peptide hormone made by the alpha cells of the
Glucagon pancreas that raises the concentration of glucose -
in the bloodstream
Antibody – large Y-shaped protein produced by Type Description
plasma cells that is used by the immune system to Found in saliva, tears, and breast milk;
identify and neutralize pathogens such as bacteria IgA prevents colonization of pathogens in
and viruses mucosal areas
Activates basophils and mast cells to
IgD
produce antimicrobial factors
Binds to allergens and triggers
Immunoglobulin IgE histamine (asthma, hayfever,
anaphylactic shock)
Provides majority of antibody-based
immunity; can cross placenta to give
IgG
passive immunity to fetus; capable of
opsonization
Eliminates pathogens in the early
IgM stages of B-cell mediated immunity
before there is sufficient IgG

27
MAJOR CLASSES OF ENZYMES
Enzyme Functions
Oxidoreductase Catalyze redox reactions
Transferase Catalyze transfer of C, N, P containing groups
Hydrolase Catalyze cleavage of bonds by addition of water
Isomerase Catalyze cleavage of C-C, C-S and certain C-N compounds
Catalyze formation of bonds between carbon and O, S, N couple to hydrolysis of high-energy
Ligase
phosphates

Properties of Enzymes
Active Site Holoenzyme vs Apoenzyme Cofactor vs Coenzyme

LOCK AND KEY THEORY VS INDUCED FIT THEORY

FACTORS AFFECTING ENZYME REACTION AND REGULATION

1 Substrate Concentration 2 Temperature – 35 to 40oC (most enzymes)
Requires that the enzyme and substrate have specific chemical groups in either an ionized or unionized state
pH in order to interact
3
Extreme pH can cause denaturation
Optimum pH – is different for different enzymes
Allosteric Binding Site Allosteric enzymes are regulated by molecules called effectors (also called modifiers)
4
that bind non-covalently at a site other than the active site
5 Covalent Modifications Either by phosphorylation or dephosphorylation
Induction and Repression Cells can also regulate the amount of enzyme present – usually by altering the rate of
6
of Enzyme Synthesis enzyme synthesis
TYPES OF ENZYME INHIBITION
Competitive Inhibition Non-competitive Inhibition
• Occurs when the inhibitor binds reversibly to the same • Occurs when the inhibitor and substrate bind at different
site that the substrate would normally occupy and sites on the enzyme
therefore competes with the substrate for that site

28
Example: statin drugs (structural analogs of the natural Example: ferrochelatase, an enzyme that catalyzes the
substrate for the enzyme HMG-CoA reductase) insertion of ferrous ion into porphyrin
*According to International Union of Biochemistry and Molecular Biology (IUBMB)
*B-Lactam antibiotics such as penicillin and amoxicillin inhibits enzymes involved in bacterial cell wall synthesis

MICHAELIS-MENTEN EQUATION
– Describes how reaction velocity varies with substrate concentration

• Km (Michaelis Constant) – characteristic of an enzyme and its particular substrate, and reflects the affinity of the
enzyme for that substrate
• Small Km = high affinity of enzyme for substrate • Large Km = low affinity of enzyme for substrate
• Rate of reaction is directly proportional to the enzyme concentration at all substrate concentration
• When substrate concentration is much less than Km, the velocity of the reaction is approximately proportional to the
substrate concentration (first order rate of reaction)
• When substrate concentration is much greater than Km, the velocity is constant and is equal to the maximum velocity
(zero-order rate of reaction)
• Lineweaver-Burk Plot – Double-reciprocal plot
– 1/Vo is plotted versus 1/[S]

CHARACTERISTICS OF CARBOHYDRATES
• The most abundant group of organic molecules in nature.
General • More than 75% of the dry weight of plants.
Statements • Less than 1% of the weight of animals is carbohydrate.
• About 65% of our diet is carbohydrate. In developing countries, figure can be up to 80%
• To serve as energy sources (also storage form of energy)
• To serve as structural component of many organisms; cell walls of bacteria, exoskeleton of many insects,
Functions fibrous cellulose of plants
• They can also serve as building blocks in the synthesis of other biomolecules (for example the nucleic
acids DNA and RNA)
Empirical • Most Carbohydrates: Cn(H2O)n or [Cn H2n On]
Formula • One molecule of water for each carbon (hydrates of carbon)
• All carbohydrates are chiral molecules (exist in a number of different orientations)
• Usually classified as D and L according to the arrangement of the OH group on the carbon furthest from
Chirality the carbonyl group.
• System is based on a comparison with the stereochemistry of the simplest carbohydrate glyceraldehyde
(C3H6O3).
• Most carbohydrates in nature have the D config. Most amino acids in nature have the L config.

29
Chirality
D-Glyceraldehyde L-Glyceraldehyde
• The D and L forms of any specific compound are Enantiomers (mirror images) of each other
Carbonyl group of carbohydrates is very reactive and can react with: (1) and (2)
1 Water to form Gem-diols

• 2 OH- groups bound to the same (geminal) carbon

• Gem-diol formation is freely reversible, although it can be a relatively slow process (seconds to
minutes), depending on the structure of the sugar.
Reactivity 2 OH- groups to form Hemi-Acetals, Acetals, Hemi-Ketals and Ketals
• Aldehyde + Alcohol → Hemiacetal + Alcohol → Acetal

• Ketone + Alcohol →Hemiketal + Alcohol →Ketal

*Hemiacetals and Hemiketals formation is reversible | **Acetals and Ketals are stable (i.e. not broken down except under extreme conditions)
MONOSACCHARIDES AND SIMPLE SUGARS
A Classification Based on the Type of Functional Group in the Sugar
• Most simple sugars contain a single carbon chain with multiple OH groups & contain either aldehyde or ketone groups
• If the sugar contains an aldehyde it is called an Aldose • If the sugar contains a ketone it is called a Ketose
Aldose

30
Ketoses

B Classification Based on Number of Carbons

Number of Carbons Type Example
3 Triose Glyceraldehyde
4 Tetrose Erythrose
5 Pentose Ribose
6 Hexose Glucose
7 Heptose Sedoheptulose
*Carbohydrates with 8 and 9 carbons are also possible

REACTIONS OF SIMPLE CARBOHYDRATES

Reaction Description
• Equilibrium exists between the open chain and the two cyclic forms of a carbohydrate
• Carbohydrates can exist as either linear (open chain) or as cyclic structures. For C5, C6 and C7
carbohydrates the cyclic form is the most common
• Fischer Projection: Open Chain Form • Haworth Projection: Cyclic Form

Mutarotation

• Pyranose: 6 membered rings with 5 carbons + 1 oxygen (more common)

• Furanose: 5 membered rings with 4 carbons + 1 oxygen
Phosphate • OH- groups of carbohydrates (like other OH groups) can react with phosphoric acids to form
Formation phosphate esters. R-OH → R-O-PO32-
Aldonic Acid
• If the aldehyde group of Carbohydrates is oxidized (CHO → COOH)
Ex: For Glucose →product is called Gluconic Acid | Ribose → Ribonic Acid

Oxidation Uronic Acid

• If the terminal OH group of Carbohydrates is oxidized ( CH2OH → COOH )
Ex: Glucose →product is called Glucuronic Acid (Vit. C is a derivative of Glucuronic Acid)

31
Aldaric Acid
• If both terminal carbon atoms to carboxyl groups are oxidized

Mutorotation

Alditols
• The aldehyde or ketone grp. of carbohydrates can be reduced to form sugar alcohols
R-CHO → R-CH2OH
Reduction • Alditols are used in processing foods and pharmaceuticals
• The Alditol of Glucose is called Sorbitol (Glucitol)
↳ It is sometimes given to diabetics.
↳ It is approximately 60% as sweet as glucose
• An OH- group of carbohydrates is often replaced by an amino group NH2
• The Amino group is sometimes acetylated NH2 → NHCO-CH3
• Glucosamine and N-Acetyl Glucosamine are amino derivatives of Glucose.
• They are important components of structural carbohydrates and are often linked to other
Amine biomolecules (proteins or lipids)
Formation

• Linking of carbohydrates together

Type No. of Sugar Units (*SU) Examples
Polymerization Monosaccharides 1 SU Ribose (5C), Glucose, Fructose, Galactose (6C)
Disaccharides 2 SU Sucrose, Maltose and Lactose
Oligosaccharides Few sugars (<10) -
Polysaccharides >2 SU Amylose, Starch, Cellulose

EPIMERS
• Two carbohydrates that differ only by the orientation of a single OH group (since carbohydrates vary by the number
and position of the OH groups)

ANOMERS
• For 5, 6 or 7 carbon carbohydrates, after cyclisation the carbonyl carbon becomes a chiral center
• Two different orientations are possible about this carbon (anomeric carbon)
– If the OH group at the anomeric carbon and the CH2OH groups are on opposite sides of the plane of
α Anomer
the ring (i.e. one above the ring and one below the ring)

32
– If the OH group at the anomeric carbon is on the same side of the ring as the CH2OH group (i.e. both
β Anomer
above or both below the plane of the ring)

EXAMPLES OF 5-CARBON SUGARS

Sugar Description
• Most common 5 carbon sugar
D-Ribose • Building block of a number of biomolecules including DNA and RNA and also molecules involved in
energy transfer such as ATP
Arabinose • Building block of several Anticancer drugs
EXAMPLES OF 6-CARBON SUGARS
• Also known as Dextrose or Blood Sugar
D-Glucose • Primary source of Carbon atoms and also energy for most tissues in the body
• Only nutrient for the brain
• Other important 6 Carbon sugars
Galactose, • Can also be converted (via phosphate derivatives) to Glucose
Mannose, • Galactosemia – genetic disease in which the enzyme responsible for galactose breakdown is
Fructose missing. Accumulation of Galactose leads to damage to the liver and brain. The most effective
treatment is early diagnosis and removal of Galactose from the diet
*Carbohydrates with 7 or more carbons are rare. The 9-carbon sugar Neuraminic acid and its N-acetyl derivative Sialic Acid is found in nerve cells.

POLYSACCHARIDES
• Most mono- and disaccharide names end in – ose
Nomenclature
• Mono and disaccharides taste sweet and are sometimes called sugars – all water soluble
• Anomeric hydroxyl group and a hydroxyl group of another sugar or some other compound can
Glycosidic Bonds join together, splitting out water.
R-OH + HO-R’ →R-O-R’ + H2O
DISACCHARIDES
Disaccharide Description
• Disaccharide with an α(1,4) glycosidic linkage between the C1 hydroxyl of one glucose and the
C4 hydroxyl of a second glucose. Maltose is the α anomer, because the O at C1 points down from
the ring.

Maltose

• Product of cellulose breakdown

• The β(1,4) glycosidic linkage is represented as a "zig-zag" line, but one glucose residue is actually
flipped over relative to the other.
Cellobiose

33
• Common table sugar
• Has a glycosidic bond linking the anomeric hydroxyls of glucose and fructose. Because the
configuration at the anomeric carbon of glucose is α(O points down from the ring), the linkage is
designated. The full name is α-D-glucopyranosyl- (1,2)- β-D- fructopyranose.

Sucrose

• Milk sugar
• Composed of glucose and galactose with β(1,4) linkage from the anomeric hydroxyl of galactose.
Its full name is β-D- galactopyranosyl-(1,4)- α-D-glucopyranose
• Most carbohydrates are produced by plants; Lactose is one of the few that is only produced by
animals

Lactose

• Lactose Intolerance – inability to digest the lactose in milk due to a deficiency of in the enzyme
Lactase. Treatment:
↳ Avoiding lactose in diet
↳ Giving patients extra Lactase
↳ Some special forms of milk have been pretreated with Lactase (or made lactose-free)
REDUCING SUGARS
• Lactose and Maltose are reducing sugars – each case one of the anomeric carbons is available for mutarotation.
• Can react with chemical reagents such as Benedict’s and Fehling’s solutions
HYDROLYSIS OF DISACCHARIDES TO MONOSACCHARIDES

34
POLYSACCHARIDES
• They have many carbohydrate units (up to 1,000,000)
• They are all non-reducing carbohydrates
General • They do not undergo mutarotation (ring opening)
Statements
• They are not sweet tasting
• They have limited solubility in water
• Storage forms of energy
Functions
• Structural Materials
Examples Description
• Main storage form of glucose in plants
• Found in many foods including rice, pasta, potatoes, wheat and cereal
• Broken down to glucose units by a group of enzymes called Amylases
• Two forms of starch: (1) and (2)
1 Amylose (20%) – which is less branched and composed mainly of α-1,4 Glycosidic Bonds (contains
250-4000 subunits arranged in a helix)

Starch

2 Amylopectin (80%) – which is similar to glycogen and has both α-1,4 and α-1,6 linkages

• Structural component of plant cells

• Used to construct rigid cell walls in plants
• Most abundant organic substance on earth
• Contains β-1,4 Glycosidic linkages

Cellulose

Cellulase
• Bacteria have enzymes that can break down cellulose. Some animals such as cows, have bacteria in
their stomach that break down cellulose
• Broken down by the enzyme Cellulase
• Humans and other animals have enzymes that can break down starch or glycogen but not cellulose
Pectins
• Soluble polysaccharides often found in fruits
Chitin
• Polymer of glucosamine which forms a very rigid polymer
• Used by many insects and other animals to form tough exoskeleton
Others Dextran
• Complex α-1,6 and α-1,3 branched glucan used medicinally as an antithrombotic agent
• Synthesized from sucrose by certain lactic acid bacteria such as Strep Mutans → dental plaque
Mucopolysaccharides
• Polysaccharides that contain either an amino sugar or a derivative of an amino sugar

35
DIETARY FIBERS
• General name given to materials from plants that the body cannot digest
• Fiber helps digestion by improving bowel function
• Increased levels of fiber in the diet lead to reductions or improvements in a variety of conditions including heart,
disease, colon cancer, and diabetes
• Pectins and Gums (soluble fibers consist of molecules)
• Cellulose (main insoluble fiber)

GLYCOPROTEINS
Carbohydrates Attached to Protein
Type of Linkage Description
N-Linked Glycoprotein • Carbohydrate attached to nitrogen in the side chain of asparagine
O-Linked Glycoprotein • Carbohydrate attached to oxygen in the side chain of serine and threonine
Non-Enzymatic • Synthetized chemically by adding sugar to polypeptides
Glycosylated Glycoprotein
Functions Examples
• Proteoglycan-linking protein
• Abundant in nerve tissues (gray matter)
• Blood clotting mechanism as prothrombin, thrombin, and fibrinogen
Structure • Rigidity of plant cells (cellulose)
• Glycocalyx of bacteria
• Flagella of bacteria
• Mucins – high molecular weight glycoproteins protecting internal epithelial surfaces (RT, GIT, UT)
Protection
• Sweat gland secretions to protect the skin
Reproduction • Important in binding of a sperm cell to the surface of the egg
• HCG (Human Chorionic Gonadotropin)
Hormone
• EPO (Erythropoietin)
Immunological • Specificity of different antibodies is determined by the carbohydrate structure in the glycoprotein
Response • B-cells and T-cells contain surface glycoproteins which can bind certain antigens

ABO BLOOD GROUP ANTIGEN

Blood Antigens Antibodies Genotype

Group Present Present

A A B AA or AO

B B A BB or BO

AB AB None AB

O None A and B OO

*The sequence of oligosaccharides determines whether the antigen is A, B, or A1

36
TESTS FOR CARBOHYDRATES
Test Positive Visible Result Reactive Group
Iodine Test Blue/Red/Purple Solution Helical Polysaccharide
Molisch Test Violet ring at junction of two layers
(Alpha-Naphthol Reaction)
Anthrone Test Green/Blue-Green color
Seliwanoff’s Test Bright Cherry-Red color (ketohexoses) Tests based on the
(Resorcinol) production of
Green solution & precipitate (pentoses) furfural or furfural
Bial’s Test Tauber’s Test for Ketoses (more specific test for pentoses, derivative
(Orcinol Test) also called Aminoguanidine Reaction; Bright Reddish-Purple
solution)
Tollen’s Phloroglucinol Test Red solution
Benedict’s Test Brick-Red precipitate
(Cupric Sulfate)
Barfoed’s Test Brick-Red precipitate (monosaccharides)
(Cupric Acetate)
Nylander Test Black precipitate (metallic bismuth is formed)
Tests based on the
(Bismuth Oxynitrate)
reducing property
Characteristic yellow crystals under the microscope
of carbohydrates
Glucosazone, Mannosazone, Fructosazone - bundles of
grass”
Phenylhydrazine Test
(Osazone Formation) Lactosazone – Powder puff-like
Maltosazone – Star-shaped
Galactosazone – Rod-shaped
Mannose – readily forms osazones – as fast as 1 minute

LIPIDS
• Chemically diverse group of compounds related more by their physical rather than
chemical properties
General Statements
• Insoluble in water (hydrophobic)
• Soluble in organic solvents like ether, benzene, chloroform, etc.
Cell Membrane Structure
• Building blocks for cell membranes
• Creates a barrier for the cell but still allows for continuous flow of materials
Thermal and Electrical Insulator
• Maintains normal temperature in the subcutaneous tissues
Functions Energy Storage
• Lipids are energy source; fats stored in adipose tissues as energy reserve
• Classified as metabolic fuel
Messengers: Hormones and Vitamins
• Hormones – communication between cells
• Vitamins – assist in the regulation of biochemical processes
Precursor Lipids
• Fatty Acids
Simple Lipids
Classifications • Fats • Waxes
Complex Lipids
• Phospholipids • Glycolipids • Others: Lipoproteins, Sulfolipids, Aminolipids
Derived Lipids
• Steroids • Others: Hormones, Vitamins, Ketone Bodies

37
FATTY ACIDS
• Occur mainly as esters in natural fats and oils, also as free fatty acids
• They are monocarboxylic acid containing hydrocarbon chains of variable length
• In natural fats they are usually:
↳ Straight chain derivatives: saturated or unsaturated
↳ Contain an even number of carbon (synthesized from 2-C units)
Types Description
• Contain no double bond
• Example: Palmitic Acid C16 Fatty Acid
Saturated
Esterification Hydrolysis
RCOOH + ROH → R-C-OR + H2O RC-C-OR +H2O → R-C-OH + ROH
Acid-Base RCOOH +NaOH → RCOO- Na+ +H2O
• Contain one or more double bonds
• Example: Oleic Acid C18 with 1 double bond at C9 (18:1Δ9)

Types of Unsaturated Fatty Acids

1 Monounsaturated (Monoenoic Acids) – contains 1 double bond
2 Polyunsaturated (Polyenoic Acids) – contains 2 or more double bonds
3 Eicosanoids – derived from Eicosa-Polyenoic Acids (C20)
Isomeric Forms of Unsaturated Fatty Acids
1 Cis-Isomer – similar or identical groups are on the same side of the double bond
Unsaturated 2 Trans-Isomer – groups are on opposite sides of a double bond
*Most FA‘s in nature are in cis-configuration
Consequence of Configuration
• Due to the shape of cis-isomer, unsaturated fatty acids don’t pack as closely as saturated fatty acids
• Unsaturated Fatty Acids – lower melting point • Saturated Fatty Acids – higher melting point
Common Reactions
• They can undergo the same reaction • Will also undergo addition
• Most common addition is hydrogenation

R-C=C-CH2CH2COOH R-CH2CH2CH2CH2COOH
• Used to convert vegetable oils to margarine during hydrogenation to produce the trans fatty acids
Type Description
• Discovered & isolated from human semen in the 1930s by Ulf von Euler of Sweden
• Like hormones they act as chemical messengers, but do not move to other sites, but
work right within the cells where they are synthesized.
• Functions: local hormone, stimulation of steroidogenesis in the ovary, inhibition of
gastric secretion, uterine contraction, lowering of BP, generation of inflammatory
Prostaglandins response
• Potent mediators of inflammation
Eicosanoids • Catalyzed by cyclooxygenase (PGH synthase)
• Cyclooxygenase (COX) is inhibited by a family of drugs known as non-steroidal anti-
inflammatory drugs or NSAIDs.
↳ Aspirin, ibuprofen, flurbiprofen and acetaminophen are all NSAIDs
• Stimulates constriction and clotting of platelets
Thromboxanes • Produced in platelets by thromboxane synthetase, which is produced from the
endoperoxides by the COX enzyme from arachidonic acid
• Synthesized in the cell from arachidonic acid by 5-lipoxygenase
Leukotrienes • Involved in asthmatic & allergic reactions and act to sustain inflammatory reactions
• Leukotriene Antagonists – used to treat asthma: Montelukast sodium & Zafirlukast

38
Eicosanoids

LIPIDS
Simple Lipids
Type Description
• Esters of fatty acids with glycerol • Oil – fat in the liquid state • Main function - energy storage

Fats
(Acylglycerols
/Glycerides)

• Esters of fatty acids with higher molecular weight monohydric alcohols

• Also functions for energy storage Fatty Acid Long Alcohol Chain
Waxes Ex: Beeswax: CH3(CH2)24COO-(CH2)29CH3 Ex: Carnauba: CH3(CH2)30COO-(CH2)33CH3
• Glands under the abdomen of bees secrete a wax, • Carnauba palm leaves, a thick coating of wax,
which they use to construct the honeycombs which can be harvested from the dried leaves
Complex Lipids
Type Description
• Lipids containing, in addition to fatty acids and an alcohol, a phosphoric acid residue
Phosphilipids
Ex: Glycerophospholipids – alcohol is glycerol Sphingophospholipids – alcohol is sphingosine
G Cardiolipin
L FATTY ACID
Y • Also called diphosphatidylglycerol
C
E FATTY ACID • 2 molecules of PA esterified through their PO4- groups to an additional molecule of glycerol
R
O Plasmalogen
L PO4 AMINO ALC
• Fatty acid is attached by an ether rather than an ester linkage at C1 of the core glycerol molecule

39
Sphingomyelin
Phospholipids • Backbone is the amino alcohol sphingosine rather than glycerol
• Important constituent of myelin of nerve fibers
• Also called glycosphingolipids
• Widely distributed in every tissue of the body particularly in the nervous tissue such as brain
• Also occurs in outer leaflet of the plasma membrane; contributes to cell surface carbohydrates
• Lipids containing a fatty acid, sphingosine and carbohydrate
Ex: Cerebrosides:(monoglycosylceramides)
↳ Major glycolipids found in animal brain and other nervous tissues
Ex: Gangliosides
↳ 1942: German scientist Ernst Klenk – lipids newly isolated from ganglion cells of brain
↳ Derived from glucosylceramide that contain in addition one or more molecules of sialic acid
Glycolipids
(neuraminic acid in human tissues)
SPHINGOSINE Leukodystrophies
FATTY ACID

• Genetically determined progressive disorders that affect the brain, spinal cord & peripheral
nerves
SUGAR ↳ Adenoleukodystrophy (ALD) – serious progressive, genetic disorder which affects the adrenal
gland and the white matter of the nervous system; abnormal peroxisomes lead to the
accumulation of very long chain fatty acids in tissues esp. the brain and adrenal glands destroying
the myelin sheath.
↳ Krabbe Disease (Globoid Cell Leukodystrophy) – inherited autosomal recessive disease caused
by deficiency of galactocerebrosidase (GALC) resulting in insufficient GALC activity to properly
metabolize certain galactolipids, including galactosylceramide and psychosine.
Tay Sachs Disease
• A fatal, recessive genetic disorder in children that causes progressive destruction of the central
nervous system
• Due to the accumulation of harmful quantities of ganglioside GM2 in the nerve cells in the brain
• Absence of the enzyme hexosaminidase A
• Occurs significantly high in persons of eastern European (Ashkenazi) Jewish descent
Derived Lipids: Steroids

• All of the steroids have a common dimethyl cyclic nucleus

• Cyclopentanophenanthrene Nucleus

• 4 rings also called gonane

STEROIDS
Type Descriptions
• With 8-10 C atoms in the side chain at C17 and α alcohol hydroxyl group at C3
• Cholesterol is major sterol in animal tissues Plant sterol: β-sitosterol poorly absorbed by humans
Cholesterol
• Synthesized mostly in the liver
Sterols • It is a constituent of animal membrane/tissues: gives rigidity and shape to the membrane
• Supplied also in the diet (meat products; chicken, beef, milk, eggs etc.) richest source is egg yolk
• It is the precursor of the steroid hormones, vitamin D and bile acids
• It can exist in the free state or binds with fatty acids at C3 to form esters: cholesteryl esters which makes
it more hydrophobic

40
Sources of Hepatic Cholesterol
• Dietary cholesterol – from chylomicron remnants • De novo synthesis
• Cholesterol from extrahepatic tissues – reverse cholesterol transport via HDL
Fats of Hepatic Cholesterol
• VLDL >> LDL – Transport to extrahepatic tissues
• Direct excretion into bile → Gallstones commonly are precipitates of cholesterol → Occurs when bile
becomes supersaturated with cholesterol (obesity, biliary stasis, infections)
• Bile acid synthesis and excretion into bile
De Novo Synthesis of Cholesterol
• Slightly less than half of the cholesterol in the body derives from biosynthesis de novo.
Primary Site: Liver Secondary Sites: Adrenal Cortex, Ovaries, Testes
Overall Eq: 18Acetyl CoA + 18ATP + 16NADPH + 4O2 → Cholesterol + 9CO2 + 16NADP + 18ADP + 18P
Drugs Used to Inhibit Cholesterol Synthesis
Sterols
• HMG-CoA Reductase Inhibitors
• Examples include various statin drugs such as Lovastatin (Mevacor), Atovarstatin (Lipitor), Simvastatin
(Zocor) and other derivatives
• Cerivastatin (Baycol) was withdrawn from the market because of increasing reports of rare, but severe
and sometimes fatal muscle damage called rhabdomyolysis
Cholesterol and Heart Disease
• High levels can accumulate and form deposits in the arteries (called plaque): blocks blood flow and may
lead to heart disease
• Condition can be controlled by diet or drugs (lipid lowering drugs: statins)
Degradation of Cholesterol
Conversion of cholesterol to bile acids, which are excreted in the feces
↳ Major excretory form of cholesterol
↳ Steroid ring is not degraded in humans
• Secretion of cholesterol into the bile, which transport it to the intestine for elimination or maybe
modified by bacteria prior to elimination forming coprostanol and cholestanol together with bile acids
make up the bulk of neutral fecal sterols
• Most common: Cholic Acid and Chenodeoxycholic Acid
• Contain 24 carbons with 2 or 3 -OH groups and a side chain that terminates in a –COOH group
Bile Acids Bile Acid Sequestrants
• Cholestyramine (Questran) binds to bile acids in the intestine. This prevents their absorption, and the
cholestyramine/bile acid complexes are eliminated in the stool. As a result, the body loses bile acids.
• Other Bile Acid Sequestrant: Colestipol (Colestid) and Colesevalam (Welchol)
• Endocrinologists classify steroid hormones into five groups of molecules, based primarily on the
receptor to which they bind
1 Glucocorticoids – cortisol is the major representative in most mammals
2 Mineralocorticoids – aldosterone being most prominent
3 Androgens – such as testosterone
Steroid 4 Estrogens – including estradiol and estrone
Hormones 5 Progestogens (Progestins) – such as progesterone
Synthesis and Secretion
• Adrenal cortex – cortisol, aldosterone
• Ovaries and Ovarian Corpus Luteum – estrogens and progestins
• Testes – testosterone
• Transcortin – transport cortisol and corticosterone
• Sex hormone-Binding Protein – transports the sex steroids (estrogens, testosterone)

41
LIPOPROTEINS
Type Description Function
High Density Picks up cholesterol from body cells and take it back
Lipoprotein Made in the liver and small intestine to the liver = “reverse cholesterol transport”
(HDL) Potential to help reverse heart disease
Made in the liver as VLDL
Low Density
Arise from VLDL once it has lost a lot of its
Lipoprotein Deliver cholesterol to all body cells
triglycerides (via lipoprotein lipase)
(LDL)
Rich in cholesterol
Very Low Made in the liver from excess dietary
Density carbohydrate and protein along with
Lipoprotein chylomicron remnant Deliver triglycerides to body cells
(VLDL) Rich in triglycerides
Made by the small intestine in the fed state
Chylomicrons Delivers triglycerides to body cells to be used as fuel
Rich in triglycerides

Difference in the Structure and Composition of Lipoproteins

Density Diameter Protein Cholesterol Phospholipid Triacylglycerol & Cholesterol
Class
(g/mL) (nm) (%) (%) (%) Ester (%)
>1.063 HDL 5-15 33 30 29 4
1.019-1.063 LDL 18-28 25 50 21 8
1.006-1.019 IDL 25-50 18 29 22 31
0.95-1.006 VLDL 30-80 10 22 18 50
<0.95 Chylomicrons 100-1000 <2 8 7 84
Bad Cholesterol: LDL, IDL, VLDL, Chylomicron, Good Cholesterol: HDL

LIPID PROFILES
• Used as part of a cardiac risk assessment to help determine an individual's risk of heart disease and to help make
decisions about what treatment may be best if there is borderline or high risk
Parameters Description
Total Cholesterol Measures all of the cholesterol in all the lipoprotein particles
High-Density Lipoprotein Measures the cholesterol in HDL particles; often called "good cholesterol" because it
Cholesterol (HDL-C) removes excess cholesterol and carries it to the liver for removal.
Calculates the cholesterol in LDL particles; often called "bad cholesterol" because it
Low-Density Lipoprotein deposits excess cholesterol in walls of blood vessels, which can contribute to
Cholesterol (HDL-C) atherosclerosis. Usually, the amount of LDL-C is calculated using the results of total
cholesterol, HDL-C, and triglycerides.
Measures all the triglycerides in all the lipoprotein particles; most is in the very low-density
Triglycerides
lipoproteins (VLDL)
Very Low-Density Lipo. Calculated from triglycerides/5; this formula is based on the typical composition of VLDL
Cholesterol (VLDL) particles
Non-HDL-C Calculated from total cholesterol minus HDL-C
Cholesterol/HDL Ratio Calculated ratio of total cholesterol to HDL-C
Low-Density Lipoprotein Included in extended profile (or advanced lipid testing)
Particle Number Measures the number of LDL particles, rather than measuring the amount of LDL-
/Concentration cholesterol
(LDL-P) May more accurately reflect heart disease risk in certain people

42
IDEAL LIPID PROFILE
Fasting Blood Level Ideal, Healthy Level
Total Cholesterol <200 mg/dl
LDL-Cholesterol <100 mg/dl
HDL-Cholesterol ≥60 mg/dl
Triglycerides <150 mg/dl

TESTS FOR LIPIDS

Test Group Positive Visible Result
Spot Test Nonvolatile Oils Semitransparent Spot
Iodine Absorption Unsaturated Lipids Purple Coloration
Acrolein Test for Glycerol, Lecithin Acrid Irritating Odor (burnt bacon)
Liebermann-Burchard Test Cholesterol Pink → Lilac → Deep-Green Color
(Acetic Anhydride-Sulfuric
Acid Test)
Salkowski Test Cholesterol Chloroform Layer: Bluish-Red/Cherry-Red Purple Sol’n.
(Sulfuric Acid Test) Acid Layer: Green Fluorescence
Molybdate Test Lecithin (but not specific) Purple Color

NUCLEIC ACID
A Classification of Nucleic Acids
• Genetic master plan of an organism is contained in the sequence of
DNA (Deoxyribonucleic Acid) deoxyribonucleotides
• Polydeoxyribonucleotide that contains deoxyribonucleotides covalently linked by
3',5'-phosphodiester bonds
• It is through the ribonucleic acid (RNA) "working copies" of the DNA that the master
plan is expressed
• Much like a single strand of DNA
• Ribonucleic acid (RNA) is like DNA except:
RNA (Ribonucleic Acid)
↳ The 2' carbon of the sugar, ribose, has a hydroxyl (OH). This sugar form is more
"fundamental" in the biochemical pathway; deoxyribose (for DNA) has to be
synthesized by removal of the 2' OH.
↳ The base uracil replaces thymine. Uracil lacks the methyl group of thymine.
Biochemically, thymine is made by methylating uracil.
B Building Blocks of Nucleic Acids

43
BASES
• Both DNA and RNA contain the same Purine Bases: adenine (A) and guanine (G)
• Both DNA and RNA contain the Pyrimidine cytosine (C)
• But they differ in their second pyrimidine base: DNA contains thymine (T) whereas contains uracil (U)

Adenine (A) R2 = H R6 = NH2 Guanine (G) R2 = NH2 R6 = O

Cytosine (C) R4 = NH2 R5 = O Uracil (U) R4 = O R5= H
Thymine (T) R4 = O R5 = CH3

NUCELEOSIDES
• Base + Sugar
• If sugar is a ribose = ribonucleoside is formed Example: Adenosine, Guanosine, Cytidine
• If sugar is a deoxyribose = deoxyribonucleoside is formed Example: deoxyadenosine, deoxyguanosine

NUCLEOTIDES
• Important regulatory compounds for pathways of intermediary metabolism, inhibiting or activating key enzymes
• Composed of a nitrogenous base, a pentose monosaccharide, and one, two or three phosphate groups
• Nitrogen-containing bases belong to two families of compounds: the purines and pyrimidines
• Mononucleotide: Monophosphate, Diphosphate, Triphosphate
• Dinucleotide – Singly esterified 5'-phosphate of a nucleotide can esterify a second alcohol functional group (-OH),
forming a diester
• Deoxynucleotide – Mono-, di-, or triphosphate esters of deoxynucleosides
↳ Phosphate group is attached by an ester linkage to the 5'-hydroxyl of the deoxyribose
↳ Ribonucleotide Reductase:
– also called ribonucleoside diphosphate reductase is specific for the reduction of nucleoside diphosphate to their deoxy-
forms
– Reduces the hydroxyl at carbon 2 of the ribose sugar in the rNDP to a hydrogen, forming a deoxyribose sugar and a
corresponding dNDP
PHOSPHODIESTER BOND
• Nucleotides are linked in the polynucleotides through the formation of a phosphodiester bond (between the 3' and
5' positions of the pentose)
• Endonuclease – cleave the chain at positions inside the chain
• Exonuclease – cleave the chain by removing individual nucleotides from one of the two ends

44
WATSON AND CRICK MODEL
• James Watson and Francis Crick inferred a structural model for DNA known as the Watson and Crick Model or the
double helix
• Two chains are coiled around a common axis
• The chains are paired in an antiparallel manner – that is, the 5’-end of one strand is paired with the 3’-end of the
other strand
• Characteristics of Double Helix:
1 Two long polynucleotide chains are coiled around a central axis, forming a right-handed double helix
2 The 2 chains are antiparallel
The bases of both chains are flat structures perpendicular to the axis – they are stacked on one another, 3.4Å
3
(0.34nm) apart and are located on the inside of the helix
4 The bases of opposite chains are paired to one another as the result of the formation of H bonds; only A=T and G≡C
5 Each complete turn of the helix is 34Å (3.4nm) long; hence 10 base pairs exist per turn in each chain
In any segment of the molecule, alternating larger major grooves and smaller minor grooves are apparent along
6
the axis
7 The double axis measures 20Å (2nm) in diameter
BASE PAIRING
• Bases of one strand are paired with the bases of the second strand, so that adenine is always paired with a thymine,
and cytosine with a guanine
• Base pairs are held together by hydrogen bonds: 2 between A and T, and 3 between G and C | (A=T and G≡C)
• Chargaff’s Principle – there is exactly as much purine (adenine and guanine) in the nucleus as there is pyrimidine
(thymine and cytosine)
SEPARATION OF STRANDS
• Two strands separate when the hydrogen bonds are disrupted between base pairs
• Causes – change in pH, heating
• Melting Temperature – temperature at which half of the helical structure is lost
• Denaturation – loss of helical structure
• Renaturation/Reannealing – reformation of the double helix by complementary strands

45
TYPES OF DNA
B-form (right-handed) A-form (right-handed)
• Hydrated form • Moderately dehydrated form
• Described by Watson and Crick Z-form (left-handed)
• Chromosomal DNA • Occurs rarely

LEVELS OF ORGANIZATION OF NUCLEIC ACIDS

Level Description
Primary Structure • Linear sequence of nucleotides
Secondary Structure • Interaction between bases
• Three-dimensional shape into which the entire chain is folded
• Differs in four structural forms:
Tertiary Structure ↳ Left of right handedness
↳ Length of the turn of helix
↳ Number of base pairs per turn
↳ Difference in size between major and minor groove
• Higher-level organization of nucleic acids
Quaternary Structure • Refers to the interactions of the nucleic acids with other molecules
• Most Common: Chromatin
CENTRAL DOGMA OF MOLECULAR BIOLOGY
“DNA makes RNA and RNA makes protein”
Flow of Genetic Information:

TYPES OF RNA
• Template for protein synthesis or translation; they carry information from the DNA
Messenger RNA (mRNA)
to the cellular protein synthetic machinery
• Carries amino acids in an active form to the ribosome for peptide bond formation, in
a sequence dictated by the mRNA template
Transfer RNA (tRNA) • Anticodon – complementary to the codon for the particular amino acid; can bind to
the codon on mRNA
• Aminoacyl Attachment Site – site at which the amino acid is attached to the tRNA
molecule
• Major component of ribosomes, plays a catalytic and a structural role in protein
Ribosomal RNA (rRNA) synthesis
• Assembly site or factory for protein synthesis involves ribosomes
TYPES OF POLYMERASES
• Synthesis & repair of polynucleotide segments is catalyzed by DNA-dependent
polymerases
DNA-Dependent DNA ↳ Polymerase I is involved in both synthesis and repair
↳ Polymerase II is also involved in DNA repair, but its function is highly specialized
↳ Polymerase III is involved in DNA synthesis
• Catalyze the formation of phosphodiester bonds from a free 3'-hydroxyl group of RNA
strand and the activated α-phosphorous atom of ribonucleoside triphosphate
• Human RNA Polymerases:
DNA-Dependent RNA ↳ RNA polymerase I present in the nucleus is responsible for transcribing the genes
for rRNA
↳ RNA polymerase II is responsible for synthesizing hnRNA, the precursor of mRNA
↳ RNA polymerase III mediates the synthesis of tRNA, 5S RNA of the ribosome, and
other small RNA molecules

46
• Also called reverse transcriptase
• Retroviruses such as human immunodeficiency virus (HIV) and human T-cell
RNA-Dependent DNA lymphotropic virus contain reverse transcriptase
• It is an enzyme that catalyzes RNA-directed DNA synthesis
• Reverse signifies that biological information flows from RNA to DNA, opposite to the
usual direction of transfer
PROTEIN SYNTHESIS
Gene Genetic Code
• Different amino acids and the order in which they are
• All genes are translated except for genes for RNA molecules
joined up determines the sort of protein being
(rRNA & tRNA)
produced
• Represent the genetic information transcribed into a single • Groups of three bases called codons control the
RNA molecule, which is in turn translated into a single protein production of a particular amino acid in the cytoplasm
Basic unit of genetic information of the cell
Basic Components of a Gene
• Base sequence (Codons)
• Promoter
• Terminator
• Transcription Start Site
• Transcription Stop Site
TRANSCRIPTION
1| Initiation 3 | Termination
•DNA is transcribed by RNA polymerase (DNA-dependent • RNA polymerase reaches terminator sequence of DNA
RNA polymerase)
• Binding of RNA polymerase to promoter region of gene • Release of mRNA
2| Elongation • Detachment of RNA polymerase from DNA
• Transcription factors unwind the DNA strand • RNA polymerase reaches terminator sequence of DNA
• Release of mRNA → • Detachment of RNA polymerase
• The antisense strand serves as the template
from DNA

47
TRANSLATION
• Attachment of the ribosome to the ribosome binding site in mRNA
• tRNA enters the ribosome at the A site
• tRNA shifts to the P site and the amino acid is added to the peptide chain
• tRNA moves to E site and is ejected from the ribosome

MUTATIONS
• Permanent change in the base sequence of DNA • Many mutations are silent mutations
• Mutant – organism with the mutated gene • Wild Type – parent organism
• Many mutations are silent mutations • Permanent change in the base sequence of DNA
• Mutagen – agent in the environment that brought about mutation
• Spontaneous Mutation – • Absence of mutagen • Occasional mistakes during DNA replication
• Induced Mutation – • Presence of mutagen
Type of Mutation Description
A Point Mutation • Base substitution
• Base substitution results to amino acid substitution in the synthesized protein
Missense Mutation
Ex: Sickle Cell Disease
• Base substitution prevents synthesis of a complete functional protein
Nonsense Mutation
• Stop codons are formed
• One of a few nucleotide pairs are deleted or inserted in the DNA
B Frameshift Mutation
• Can shift the translational reading frame
TYPES OF MUTAGENS
Physical Mutagens DNA-Reactive Chemicals Intercalating Agents Metals Biological Agents
• Reactive Oxygen
• Ionizing • Arsenic • Transposon
• Species (ROS) • Ethidium Bromide
Radiations
• Nitrous Acid • Mercury
• Bacteria (H. pylori)
• Bromine • Cadmium
• Ultraviolet • Benzene • Proflavine
• Nickel • Virus
• Psoralen
Mutagens – may be of physical, chemical or biological origin. They may act directly on the DNA, causing direct damage
to the DNA, and most often result in replication error.
RECOMBINANT DNA TECHNOLOGY
• Purposeful manipulation of genetic material to alter the characteristics of an organism in a desired way

48
BIOTECHNOLOGY
• Use of microorganisms, cells, or cell components to make a product
• Vaccines • Interferons • Monoclonal Antibodies • Blood Factors •Hormones
POLYMERASE CHAIN REACTION (PCR)
• Technique by which small samples of DNA can be quickly amplified for analysis
• Principle:
↳ When DNA is heated, the strands separate or “melt”
↳ If the single stranded sequence can be copied by DNA polymerase, the original sequence is duplicated
↳ If the process is repeated many times, there is an exponential increase in the number of copies
STEPS IN POLYMERASE CHAIN REACTION
Steps Description
Denaturation Strands of DNA are melted apart by heating to 95°C
Annealing Temperature is reduced to 55°C to allow primers to anneal to the target DNA
Temperature is changed to the optimum temperature (72°C) in order for the DNA
Polymerization
polymerase to catalyze extension of the primers

GENERATION AND STORAGE OF METABOLIC ENZYMES

METABOLISM
• Set of chemical reactions that occur in a cell, which enable it to keep living, growing and dividing. Metabolic processes
are usually classified as:
• Catabolism – obtaining energy and reducing power from nutrients
• Anabolism – production of new cell components, usually through processes that require energy and reducing power
obtained from nutrient catabolism
Pathway Description
Glycolysis Glucose oxidation in order to obtain ATP
Kreb’s Cycle Acetyl-CoA oxidation in order to obtain GTP and valuable intermediates
B-Oxidation of Fatty Acids Fatty acids breakdown into Acetyl-CoA, to be used for Kreb’s Cycle
Pentose Phosphate Synthesis of pentoses and release of the reducing power needed for anabolic reactions
STORAGE OF METABOLIC FAT
Pathway Description
Glycogenesis Glycogen formation from glucose
Gluconeogenesis Glucose synthesis from smaller precursors, to be used by the brain
Lipogenesis Acetyl-CoA conversion to fatty acids

49
ENZYMATIC CONTROL OF METABOLIC PATHWAYS
Pathway Regulation Regulator
• Can be regulated at three key points:
• Hexokinase – is inhibited by glucose-6-P (product inhibition)
• Phosphofructokinase – is inhibited by ATP and citrate (which signals the abundance of
citric acid cycle intermediates). It is also inhibited by H+, which becomes important under
Glycolysis anaerobiosis (lactic fermentation produces lactic acid, resulting on a lowering of the pH).
Probably this mechanism prevents the cell from using all its ATP stock in the
phosphofructokinase reaction, which would prevent glucose activation by hexokinase. It
is stimulated by its substrate (fructose-6-phosphate), AMP and ADP (which signal the lack
of available energy), etc.
• Pyruvate Kinase – inhibited by ATP and acetyl-CoA
• Regulated mostly by substrate availability, product inhibition and by some cycle
intermediates:
• Pyruvate Dehydrogenase – is inhibited by its products, Acetyl-CoA and NADH
• Citrate Synthase – inhibited by its product, citrate. Also inhibited by NADH & Succinyl
Kreb’s Cycle • Isocitrate Dehydrogenase and A-Ketoglutarate Dehydrogenase – like citrate synthase,
these are inhibited by NADH and Succinyl-CoA. Isocitrate dehydrogenase is also inhibited
by ATP and stimulated by ADP. All aforementioned dehydrogenases are stimulated by
Ca2+. This makes sense in the muscle, since Ca2+ release from the sarcoplasmic reticulum
triggers muscle contraction, which requires a lot of energy. This way, the same "second
messenger" activates an energy-demanding task and the means to produce that energy.
• Acyl-CoA movement into the mitochondrion is a crucial factor in regulation. Malonyl-
CoA (which is present in the cytoplasm in high amounts when metabolic fuels are
Fatty Acids Metabolism abundant) inhibits carnitine acyltransferase, thereby preventing acyl-CoA from entering
the mitochondrion. Furthermore, 3-hydroxyacyl-CoA dehydrogenase is inhibited by
NADH and thiolase is inhibited by acetyl-CoA, so that fatty acids will not be oxidized when
there are plenty of energy-yielding substrates in the cell.
• Metabolic flow through the pentose phosphate pathway is controlled by the activity of
Pentose Phosphate
glucose-6-phosphate dehydrogenase, which is controlled by NADP+ availability
• Flow is regulated in the gluconeogenesis-specific reactions. Pyruvate carboxilase is
Gluconeogenisis activated by acetyl-CoA, which signals the abundance of citric acid cycle intermediates,
i.e., a decreased need of glucose

ROLE OF HORMONES
• Mainly effected through the action of two hormones synthesized by the pancreas: insulin and glucagon
• Insulin is released by the pancreas when blood glucose levels are high, i.e., after a meal
↳ Stimulates glucose uptake by the muscle, glycogen synthesis, and triacyl-glyceride synthesis by the adipose tissue
↳ Inhibits gluconeogenesis and glycogen degradation
• Glucagon is released by pancreas when blood glucose levels drop too much
↳ Its effects are opposite those of insulin: in liver, glucagon stimulates glycogen degradation and the absorption of
gluconeogenic amino acids
↳ Inhibits glycogen synthesis and promotes the release of fatty acids by adipose tissue

50
51
GLYCOLYSIS

Tricarboxylic Acid Cycle B-Oxidation of Fatty Acids Pentose Phosphate

(Citric Acid Cycle/Kreb’s Cycle)

52
STORAGE OF METABOLIC FATS
Glycogenesis Gluconeogenesis Mono/Disaccharide Metabolism

LIPOGENESIS CODON CHART

53