Protein

Cellular Function of Proteins

 Enzymes are biological catalyst. Example: Pepsin, Trypsin

 Defense Proteins include antibodies (Immunoglobulins) which are specific protein

molecules produced by specialized cells of the immune system in response to foreign
antigens.
 Transport Protein carries material from place to another in the body. Example:
transferrin, Hemoglobin and Myoglobin.
 Regulatory Protein controls many aspects of cell function, including metabolism and
reproduction. Example: Insulin and Glucagon
 Structural proteins provide mechanical support to large animals and provide them with
their outer coverings. Example: Keratin
 Movement Proteins are necessary for all forms of movement. Example: Actin and
Myosin, Flagella of sperm cell.
 Nutrient Protein serves as source of Amino acids for embroyos or infants. Example:
Albumin and Casein
 Protein structure

Primary Structure of Proteins

 It is the amino acid sequence of the protein chain.

 This structure will determine its biological active form.
 It results from the covalent bonding between the amino acids in the chain
 The amino acid sequence of a protein is encoded in DNA. Proteins are synthesized by a
series of steps called transcription (the use of a DNA strand to make a complimentary
messenger RNA strand - mRNA) and translation (the mRNA sequence is used as a
template to guide the synthesis of the chain of amino acids which make up the protein).
 Genes can change by the process of mutation during the course of evolution.
 A mutation in gene can result in a change in the primary acid sequence of a protein.

Secondary Structure of Proteins

 This level of structure describes the local folding pattern of the polypeptide backbone
and is stabilized by hydrogen bonds between N-H and C=O groups. The most common
are the orderly repeating forms known as the helix and the b sheet.
 The secondary structure is the result of hydrogen bonding between the amide
Hydrogen and carbonyl oxygen’s of the peptide bonds.

Types of Secondary Structure

 α-Helix and β-Pleated Sheet

α-Helix

 The most common type of Secondary structure. This may it be in coiled of helical
conformation.

Special Feature

 Every amide hydrogen and carbonyl oxygen is associated with the peptide backbone is
involved in a hydrogen bond when the chain coils into a α-Helix.
 Every carbonyl oxygen is hydrogen bonded to an amide hydrogen four amino acids away
in the chain.

Structural Property of a-helix

 It has great mechanical strength and is applied very efficiently in both the fibrous
protein of skin and those of muscle.
Proteins having a α-Helix structure

 Fibrous Proteins – are structural proteins arranged in fibers or sheets that have only
one type of secondary structure.

α-Keratins

 Are fibrous proteins that form the covering (hair, nails and fur) of most land animals.

β-Pleated Sheet

 The second common secondary structure in proteins resembles the pleated folds of
drapery
 All the carbonyl oxygen and amide hydrogens in a B-pleated sheet are involved in
hydrogen bonds, and the polypeptide chain is nearly completely extended.

2 Orientation of B-Pleated form

1. Parallel
 Beta sheets are parallel if the polypeptide strands run in the same direction, N-
terminus to C-terminus. The N-terminus of one beta strand will be opposite the N-
terminus of the other beta strand.
 The parallel arrangement is less stable because the geometry of the individual amino
acid molecules forces the hydrogen bonds to occur at an angle, making them longer and
thus weaker.
2. Anti-parallel
 Beta sheets are anti-parallel if the polypeptide strands run in opposite directions. The
N-terminus of one beta strand will be opposite the C-terminus of the other beta strand.
 In the anti-parallel arrangement the hydrogen bonds are aligned directly opposite each
other, making for stronger and more stable bonds.
 An anti-parallel beta-pleated sheet forms when a polypeptide chain sharply reverses
direction. This can occur in the presence of two consecutive proline residues, which
create an angled kink in the polypeptide chain and bend it back upon itself.

Silk Fibroin

 A protein whose structure is an antiparallel B-pleated sheet.

 The polypeptide chains of a B-pleated sheet are almost completely extended, and silk
does not stretch easily.
 Glycine accounts for nearly half of the amino acids of silk fibroin. Alanine and serine
account for most of the others,
Tertiary Structure of Proteins

 Tertiary structure is the complete three-dimensional (3-D) structure of a polypeptide. It

is formed spontaneously and stabilized both by side chain interactions and, in
extracellular proteins, by disulfide bonds. This folding brings distant sequences in a
linear polypeptide together into a stable structure

The structure is maintained by the following molecular interactions:

 Van der Waals Forces between the R groups of non-polar amino acids that are
hydrophobic.
 Hydrogen bonds between the polar R group of the polar amino acids
 Ionic bonds (salt bridges) between the R groups of oppositely charged amino acids
 Covalent bonds between the thiol-containing amino acids.

 Globular proteins generally have a more compact and rounded shape and have
functional roles (they do something)

Quaternary Structure

 Some proteins are made up of multiple polypeptide chains, also known as subunits.
When these subunits come together, they give the protein its quaternary structure.
 The forces that hold the quaternary structure of a protein are the same as those that
hold the tertiary structure.
 Prosthetic group – when a non-protein group is added to the functional protein.
Example: Glycoprotein

Protein digestion

 Is the degradation of protein by cellular enzymes in a process called hydrolysis.

 The macromolecules are the proteins or polypeptides themselves, and the subunits are
the amino acids.
 It takes place in two different phases:
 In the stomach, and In the small intestine
 Both of these phases of digestion are based on several types of enzymes that are called
Proteinases and Proteases.
 Proteases – endo and exo peptidases
 Enzymes that degrade proteins by hydrolysis of peptide bonds
 Proteinases- endo peptidases; proteases that show specificity for intact proteins
Protein Digestion in Mouth and Salivary Glands

 Chewing and crushing rich foods and mix them with saliva to be swallowed.

Protein Digestion in Stomach

 It is the start of protein digestion.

 Gastrin – Stimulates Parietal cells to secrete HCl; Chief cells of the gastric glands to
secrete pepsinogen
 HCl / Hydrochloric Acid – Denatures protein structure. Activates pepsinogen (Zymogen)
to pepsin.
 Pepsin- Hydrolyses proteins to smaller polypeptides and some free amino acids

Protein Digestion in Intestine

 The remainder of protein digestion occur in the small intestine as the result of the
action of enzymes such as trypsin (secreted by the pancrease) and petidases (located in
the cells that line the small intestine).
 Secretin – Stimulates the pancreas to secrete bicarbonate into the small intestine to
neutralize the gastric HCl
 Cholecystokinin- Stimulates secretion of several pancreatic enzymes with activity
optima pH 7 to 8.
 Trypsin - Activates chymotrpsinogen  chymotrypsin
 Further hydrolyze the peptides that were produced by pepsin in the stomach specifically
the peptide bonds next to Lys and Arg.
 Chymotrypsin - Cleaves peptide bonds next to Phe, Tyr, Trp, Met, Asp and His

Denaturation of Protein

 Occur when the organized structures of a globular protein, the a-helix, the B-pleated
sheet and tertiary folds become completely disorganized. However, it does not alter the
primary structure.

Factors that cause Denaturation

Temperature

 As the temperature increase, molecular movement also increase and the bonds within
the cells vibrate more violently which results to disruption of protein structure.
 Coagulation- occurs as the protein molecules unfold and become entangled. At this
point, they are no longer in solution; they have aggregated to become a solid.
  Many of the proteins in our cells is in viscous solution within our cytoplasm. To continue
to function properly they must remain in solution and maintain the correct three-
dimensional configuration.

pH

 A high concentration of hydrogen ions (low pH) will result in more groups being
protonated. Carboxyl groups (aspartic acid, glutamic acid, the carboxy terminus) and
phenolic groups are uncharged when protonated. The nitrogen groups (amines on
lysine, guanidino of arginine, and imidazole in histidine, etc.) are charged when
protonated

Organic Solvents

 Polar organic solvents, such as rubbing alcohol (2-propanol), denatured proteins by

disrupting hydrogen bonds within the protein, in addition to forming hydrogen bonds
with the solvent, water
 Nonpolar regions of these solvents interfere with hydrophobic interactions in the
interior of the protein molecules, thereby disrupting the conformation.

Heavy metals

 Mercury (Hg2+) or Lead (Pb2+) may form negatively charged side chain groups.
 Heavy metals may also bind to sulfhydryl groups of a protein that can accompanied by
loss of function.

Detergents

 Detergent have hydrophobic region and hydrophilic. When detergents interact with
proteins, they disrupt hydrophobic interactions, causing the protein chain to unfold.

Mechanical Stress

 Stirring, whipping, or shaking can disrupt the weak interactions that maintain protein
conformation. This is the reason that whipping egg whites produces a stiff meringue.
 Enzyme: Part II

Enzyme

 Greek word en, means “in”, and zyme, which means “yeast” an organic compound that
act as a catalyst for a
 Biochemical reaction; increases rate of reaction but are not changed in the process
 Water-soluble, globular proteins

Features:

a) Enhance reaction rates (pH 7.4 and 37°C). An enzyme catalyzed reaction can be 106 to
1012 times faster than unanalyzed reaction.
b) Enzymes are very specific.

Simple structure of enzyme

1. Simple enzyme – an enzyme composed only of protein.

2. Conjugated enzyme – an enzyme that has a no protein part in addition to the protein
part.
 Apoenzyme is the protein part of a conjugated enzyme cofactor is the non-
protein part of a conjugated enzyme.
 Holoenzyme is the biochemically active conjugated enzyme produced from an
apoenzyme and a cofactor.
 Apoenzyme + cofactor = holoenzyme

Why do apoenzymes need cofactors?

 A cofactor is a non-protein chemical compound that is required for the protein's

biological activity.
 Cofactors can be considered "helper molecules" that assist enzymes in their action.
Cofactors can be ions or organic molecules (called coenzymes).
 Coenzyme is a small organic molecule that serves as a cofactor in a conjugated
enzyme.
 Inorganic ion cofactors include zinc, magnesium, manganese, and iron; chloride
occasionally acts as a factor.

Role of Metal Ions in Enzymes

 Activity of many enzymes depends on the presence of potassium, magnesium, calcium,

zinc and copper ions.
 Metal activated enzymes

 Metals form a loose and easily dissociable complex.

 The metal ion can be removed by dialysis or any other such method from the enzyme
without causing any denaturation.

Metallo-enzymes

 It is bound tightly to the enzyme and is not dissociated even after several extensive
steps of purification of apoenzyme.

Metal play a variety roles such as

a) Help in either maintaining or producing (or both) active structural conformation of the
enzyme.
b) Formation of enzyme-substrate complex.
c) Making structural changes in substrate molecule.
d) Accept or donate electrons
e) Activating or functioning as nucleophiles
f) Formation of tertiary complexes with enzyme or substrate.

Nomenclature and Classification of Enzymes

 Naming provides the function of the enzyme rather than the structure; type of
reaction catalyzed and substrate are the focal points for nomenclature.
 Substrate is the reactant in an enzyme-catalyzed reaction; it is the substance upon
which the enzyme “acts”
1. Suffix –ase identifies an enzyme (urease, lipase); suffix –in in some names of
digestive enzymes (trypsin, chymotrypsin, and pepsin)
2. The type of reaction catalyzed by an enzyme is often noted with a prefix. (Oxidase,
hydrolase)
3. The identity of the substrate is often noted in addition to the type of reaction.
(glucose oxidase, pyruvate carboxylase, and succinate dehydrogenase)
 Few enzymes exist in their inactive form called proenzymes or zymogens.
 Zymogens become active after prior modification in its structure by certain agents.
Many times the active form of enzyme acts on zymogen and catalyzes its conversion
into active form – process called autocatalysis
 Properties of enzyme

A. Active sites
 Special pocket or cleft within the enzyme molecule contains amino acids side chains that
participate in substrate binding and catalysis
B. Catalytic efficiency
 Highly efficient (103- 108) times faster than uncatalyzed reactions
C. Specificity
 Highly specific, interacting only with one or a few substrates and catalyzing only one
type of chemical reaction
D. Holoenzymes
 Some enzymes require molecules other than proteins for enzymatic activity
 Holoenzymes refer to the active enzymes with its non-protein component, whereas the
enzyme without its non-protein moiety is termed an apoenzyme and is inactive.
 If the non-protein is a metal ion (Zn2+ or Fe2+ ), it is called a cofactor
 If it is a small organic molecule, it is termed as coenzyme.
 Coenzymes that only transiently associate with the enzyme are called cosubstrate
 Cosubstrates dissociate from the enzyme in an altered state. (ex. NAD+)
 Coenzymes frequently are derived from vitamins. (ex. NAD+ contains niacin and FAD
contains riboflavin)
E. E. Regulation
 Enzyme activity can be regulated, that is, increased or decreased, so that the rate of
product formation responds to cellular need.
F. Location within the cell
 Many enzymes are localized in specific organelles within the cell. Such
compartmentalization serves to isolate the reaction substrate or product from other
competing reactions. This provides a favorable environment for the reaction, and
organizes the thousands of enzymes present in the cell into purposeful pathways

Classes of Enzymes (Based on the Type of Reaction)

1. Oxidoreductase
 Requires a coenzyme that is oxidized or reduced as the substrate is reduced or oxidized.
 Lactate dehydrogenase is an oxidoreductase that removes hydrogen atoms from a
molecule.
 An organic oxidation reaction is an oxidation that increases the number of C-O
bonds and/or decreases the number of C-H bonds.
 An organic reduction reaction is a reduction that decreases the number of C-O
bonds and/or increases the number of C-H bonds.
2. Transferase
 An enzyme that catalyzes the transfer of a functional group from one molecule to
another.

Two subtypes

 Transaminase catalyzes the transfer of an amino group from one molecule to another;
 Kinases - catalyze the transfer of a phosphate group from adenosine triphosphate (ATP)
to give adenosine diphosphate (ADP) and a phosphorylated product (a product
containing an additional phosphate group)
3. Hydrolase
 An enzyme that catalyzes a hydrolysis reaction in which the addition of a water
molecule to a bond causes the bond to break. Hydrolysis is central to the process of
digestion.

4. Lyase
 It is an enzyme that catalyzes the addition of a group to a double bond or the removal of
a group to form a double bond in a manner that does not involve hydrolysis or
oxidation.
 Dehydratase – effects the removal of the components of water from a double
bond
 Hydratase – effects the addition of the components of water to a double bond
5. Isomerase
 An enzyme that catalyzes the isomerization (rearrangement of atoms) of a substrate in a
reaction, converting it into a molecule isomeric with itself.
 There is only one reactant and one product in reactions where isomerases are
operative.
6. Ligase
 It is an enzyme that catalyzes the bonding together of two molecules into one with the
participation of ATP.
 Main classes and subclasses of enzyme

 Main Classes - Selected Subclasses - Type of Reaction Catalyzed

I. Oxidoreductases - Oxidases - Oxidation of a substrate
- Reductases- Reduction of a substrate
- Dehydrogenases - Introduction of double bond (oxidation) by
formal removal of two H atoms from a substrate, with one H
being accepted by a coenzyme
II. Transferases - Transaminases - Transfer of an amino group between substrate
- Kinases - Transfer of a phosphate group between substrate
III. Hydrolases – Lipases - Hydrolysis of ester linkages in lipids
- Proteases - Hydrolysis of amide linkage in proteins
- Nucleases - Hydrolysis of sugar-phosphate ester bonds in nucleic acids
- Carbohydrases - Hydrolysis of glycosidic bonds in carbohydrates
- Phosphatases - Hydrolysis of phosphate-ester bonds
IV. Lyases – Dehydratases - Removal of H2O from a substrate
- Decarboxylases - Removal of CO2 from a substrate
- Deaminases - Removal of NH3 from a substrate
- Hydratases - Addition of H2O to a substrate
V. Isomerase - Racemases - Conversion of D-isomer to L-isomer or vice versa
- Mutases - Tranfer of a functional group from one position to another
in the same molecule
VI. Ligases - Synthetases - Formation of a new bond between two substrates, with
participation of ATP
- Carboxylases - Formation of a new bond between substrate and CO2
with participation of ATP.

Lipids

Lipids

 Actually refers to a collection of organic molecules of varying chemical composition

 Group together on the basis of their solubility in nonpolar solvents

Importance of Lipids in biological processes

 Energy Source – When oxidized, each gram of fats releases 9kcal of energy, or more
than twice the energy released by oxidation of a gram of carbohydrates.
 Energy storage – Stored in fat called adipocytes
 Cell membrane structural components – These membranes control the flow of
molecules into and out of the cells and allow cell to cell communication.
  Hormones- steroid hormones are critical chemical messengers that allow tissues to the
body to communicate with one another.
 Vitamins – The lipid-soluble vitamins; A,D,E and K, play a major role in the regulation of
several critical biological processes, including blood clotting and vision.
 Vitamin absorption – Dietary fat serves as carrier of the lipids-soluble vitamins. All are
transported into cells of the small intestine in association with fat molecules.
 Protection – Fats serves as shock absorber, or protective layer, for the vital organs
 Insulation – Fat stored beneath the skin serves to insulate the body from extremes of
cold temperature.

Fatty acid

 Are long-chain monocarboxylic acids

 Generally contain an even number of carbon atoms
 CH3(CH2) nCOOH in which n in biological system is an even integer

Saturated fatty acids

 The carbon in the chain is bonded to the maximum number of hydrogen atoms

Unsaturated Fatty Acids

 An unsaturated fat is a fat or fatty acid in which there is one or more double bond in
the fatty acid chain.

Chemical Reaction of Fatty Acids

 Identical to those of short-chain carboxylic acids

 Undergo saponification, Acid hydrolysis of esters, esterification and addition of the
double bond.

A. Esterification

 Fatty acids react with alcohol to form esters and water.

B. Acid Hydrolysis

 The reverse of esterification, producing fatty acids from esters:

C. Saponification

 It is the base-catalyzed hydrolysis of an ester. The product which is an ionized salt is

soap.
D. Reaction at the double bond (unsaturated fatty acid)

 Hydrogenation is an example of an addition reaction

Essential fatty acid

 An unsaturated fatty acid that is essential to human health, but cannot be manufactured
in the body. Abbreviated EFA.
 There are three types of EFAs: arachidonic acid, linoleic acid, and linolenic acid

Linoleic acid

 It is known as an omega-6 fatty acid, occurring widely in plant glycosides

 Linoleic acid is an essential fatty acid in human nutrition because it cannot be
synthesized by humans.
 It is used in the biosynthesis of prostaglandins (via arachidonic acid) and cell
membranes.
 Called omega-3 fatty acids. Alpha-linolenic acid, in particular, is not synthesized by
mammals and therefore is an essential dietary requirement for all mammals. Certain
nuts and vegetable oils are particularly rich in alphalinolenic acid (ALA).
 The precursor of many prostaglandins
 Precursor to prostaglandins that have inflammatory effects

EICOSANOIDS: Prostaglandins, Leukotrienes and Thromboxines

 It is a signaling molecules made by oxidation of twentycarbon essential fatty acids,

(EFAs).

3 types of Eicosanoids

 Prostaglandins, Thromboxanes, Leukotrienes

Biomedical importance of Eicosanoids

1. Blood Clotting

 Thromboxane A2 is produced by platelets in the blood and stimulates constriction of

the blood vessels and aggregation of the platelets.

2. The inflammatory response

 Prostaglandins are thought to promote certain aspects of the inflammatory response,

especially pain and fever.
3.Reproductive System

 PGE2 stimulates smooth muscle contraction, particularly uterine contractions

 2 of prostaglandins is responsible for dysmenorrhea (painful menstruation)

4. Gastrointestinal tract

 Prostaglandins have been shown to both inhibit the secretion of acid and increase the
secretion of a protective mucus layer into the stomach.

5.Kidney

 Prostaglandins produced in the kidneys cause the renal blood vessels to dilate. The
greater flow of blood through the kidney results in increased water and electrolyte
excretion.

Arachidonic acid

 It is fatty acids
 20 carbon fatty acids consisting of 4 double bonds
 Present in cell membrane and released from it during normal metabolic processes, cell
death or injury.

Prostaglandins

 Has hormone like activity and isolated from seminal fluid produced in prostate glands
 Linoleic acid – the dietary precursor of Prostaglandins
 Arachidonic acids – The immediate precursor of the main prostaglandins in human
 Prostaglandins are made at sites of tissue damage or infection, where they cause
inflammation, pain and fever as part of the healing process.
 When a blood vessel is injured, a prostaglandin called thromboxane stimulates the
formation of a blood clot to try to heal the damage; it also causes the muscle in the
blood vessel wall to contract (causing the blood vessel to narrow) to try to prevent
blood loss.
 Prostaglandins are known to regulate the female reproductive system, and are involved
in the control of ovulation, the menstrual cycle and the induction of labor.

Leukotrienes

 Leukotrienes (LTs) are most commonly found in leukocytes, mast cells, platelets, and
vascular tissues of the lung and heart.
 Leukotrienes are involved in the pathogenesis of both acute and chronic asthma. They
cause increased bronchoconstriction, mucous secretion, and vascular permeability.
Thromboxane

 It is produced in platelets by thromboxane synthetase, which is produced from the

endoperoxides by the cyclooxygenase (COX) enzyme from arachidonic acid.

Type of Thromboxane

 Thromboxane A2 (TXA2) – It is a type of thromboxane that is produced by activated

platelets and has prothrombotic properties.
 Thromboxane B2 (TXB2) – It is an inactive metabolite/ product of thromboxane A2. It
is almost completely cleared in the urine.

Glycerides

 It is lipid esters that contain the glycerol molecule and fatty acids.

Subdivided into two classes

1. Neutral glycerides
2. Phosphoglyceride

Neutral Glycerides

 Are ionic and nonpolar

 Produce by esterification of fatty acids.
 Esterification may occur at one, two or all tree position, producing monoglycerides,
diglycerides and triglycerides.
 There are no – or + charge on these molecules. These long molecules readily stack with
one another and constitute the majority of the lipids stored in the body’s fat cells.

Function of neutral glycerides in biochemical systems

 It is the storage of energy

 If more energy-rich nutrients are consumed than are required for metabolic processes,
much of the excess is converted to neutral glycerides and stored as triglycerides in fat
cells of adipose tissue.
 When energy is needed, the triglycerides are metabolized by the body, and energy is
released.

Phosphoglycerides

 Phospholipids- are a group of lipids that are phosphate esters

 Phoshoryl group makes the molecule polar and the fatty acid chain is the non-polar tail
  Known as glycerol-3-phosphate
 The most abundant membrane lipids
 Contains acyl group derived from long-chain fatty acids at C-1 and C-2 of glycerol -3-
phosphate. At C-3 the phosphoryl group is joined to glycerol by a phosphatester bonds.

Phosphatidate

 The simplest phosphoglycerides which contains free phosphoryl group

 ‘When phosphoryl group is attaching to another hydrophilic molecule, a more complex
phosphoglycerides is formed.
 Example: lecithin and cephalin

Lecithin or Phosphatidylcholine

 An amphipathic molecule
 Ionic head – is hydrophilic and interacts to water
 Nonpolar tail hydrophobic and interacts with nonpolar molecules

Cephalin or Phosphatidylethanolamine

 A phospholipid found particularly in the cells of nervous tissue; it is also the primary
phospholipid in bacteria.