Honours / Core Course (CC)

 
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BIOCHEMISTRY
Fundamentals of Biochemistry
CARBOHYDRATES
Structure and Biological importance:

Monosaccharides

Q.What is mutarotation?

As monosaccharides contain asymmetric C-atoms (e.g; two anomers of D-glucose), they may remain in various isomeric forms (D or L forms, D=dextrorotatory and L= Laevorotatory).When a monopolarized light is pass through a solution of such carbohydrates, the plane of the light is rotated certain degree to either right or left hand. Carbohydrate which can rotate light in right-hand (clock wise) direction is dextrorotatory (D-form) and that in left hand (anti clock wise ) direction is called Laevorotatoryb m ,yut (L- form) .

e.g ; the specific optical rotation values of α-D-Glucose and β-D-Glucose are +112.20 and 18.70 respectively.

However on standing, the specific optical rotation slowly changes until it reaches an equilibrium value of +52.70.This phenomenon in change of optical rotation is called mutarotation.

N.B:In Fructose ,the mutarotation value is -1130 to -920.

Disaccharides

Polysaccharides

Q.Distinguish between starch and glycogen.

(A) Starch:

Consists of 2 components—(alpha-amylose (15-35%)and alpha-amylopectin (65-85%). In alpha-amylopectin branching points occur every 24-30 glucose residues.

i)It is plant polysaccharide, stored in grains, legumes, tubers like potato and roots like carrot.

ii)Amylose and amylopectin molecules form respectively blue and violet complexes with iodine.

iiii)Starch is nonreducing, because the free anomeric OH groups of most glucose residues remain bound in glycosidic bonds.

(B) Glycogen:

Resembles that of amylopectin, but glycogen is more highly branched with branching points occurring every 8-12 glucose residues.

i)This is a type of food-storage homoglycan, stored largely in liver and muscle cells of animals and also in fungi and Saccharomyces.

ii)This molecule forms a colloidal solution in water, is dextrorotatory, gives a red-violet colour with iodine, has no reducing property and is hydrolysed by boiling with dilute mineral acids into many molecules of glucopyranose.

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Derivatives of Monosaccharides

Carbohydrate metabolism :

Glycolysis

Citric acid cycle

Pentose phosphate pathway

Q.What is pentose phosphate pathway?

This is an oxidative pathway (also called hexose monophosphate pathway shunt or the phosphogluconate pathway) that operates in the cytosol of cells. This biochemical pathway is active in tissues such as the liver, adrenal cortex, testis, adipose tissue and lactating mammary glands, where fatty acids, steroids or pentoses are synthesized.

The first part of this pathway constitutes its oxidative phase which oxidizes glucose-6-phosphate into ribulose-5-phosphate with the help of NADP+ and reduces the latter to NADPH. This is followed by the nonoxidative phase which converts ribulose 5-phosphate to ribose 5-phosphate and also changes the latter and other C5 and C4 sugars into fructose 6-phosphate and glyceraldehydes 3-phosphate.

Gluconeogenesis

Q.What is the significance of gluconeogenesis?

It maintains the liver glycogen and a steady basal level of the blood sugar during starvation, for supplying glucose to the brain, kidneys, erythrocytes and muscles. It removes from blood the metabolites of glycolysis and lipolysis and recycles them as carbohydrates. Lactate produced by glycolysis is largely released by muscles into the blood, carried to the liver and reconverted by glycogenesis to glucose which is carried to muscles for glycolysis again.

LIPIDS
Structure and Significance:

Physiologically important saturated and unsaturated fatty acids

Triacylglycerols

Q.What do you mean by rancidity effect of lipid?

Rancidity is the offensive odour and taste of fats and oils on standing exposed to air, moisture and warm temperature. This may be result from the formation of obnoxious aldehydes due to the oxidation of unsaturated traacylglycerols at the double bonds of their fatty acid residues by atmospheric oxygen and ozone (oxidative rancidification).

Rancidity may also result from the hydrolysis of ester bonds I fat molecules by enzymes of contaminating microbes and the consequent release of free fatty acids (hydrolytic rancidification).

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Phospholipids
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Q.What is lecithin?

Lecithin or phosphatidylcholines is an example of phospholipid where the nitrogen base choline bound to the phosphate group. The molecule is made of a glycerol, two fatty acids, a phosphate and a choline. In the polar head group consisting of phosphocholine, the phosphate and choline residues bear anionic and cationic charges respectively, depending on the pH. Lecithins occur in many tissues such as brain, liver, cardiac muscle and blood. Lecithin is a fat that is essential in the cells of the body. It can be found in many foods, including soybeans and egg yolks. Lecithin is taken as a medicine and is also used in the manufacturing of medicines. Lecithin is used for treating memory disorders such as dementia and Alzheimer's disease. It is the most abundant phospholipid in the cell membrane. It acts as a lipotropic factor preventing accumulation of lipid in the liver.

Sphingolipid
Glycolipids
Steroids
Eicosanoids and terpinoids
Lipid metabolism:
β-oxidation of fatty acids-
►(a).Palmitic acid {saturated (C 16:0)}

Q.State the activation phase of fattyacid prior to β-oxidation.

Before the transfer into mitochondria, fatty acids need to be changed by fatty acid thiokinases (acyl-CoA synthases) into active intermediates called acyl-CoA or fatty acid- CoA thioesters. Thiokinases are C—S ligases occurring on the outer mitochondrial membrane and in the mitochondrial matrix. A thiokinase uses ATP for adenylating the fatty acid into an enzyme-bound acyl adenylate with the release of pyrophosphate.

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►(b).Linoleicacid {unsaturated (C 18:2)}
Fatty acid biosynthesis
PROTEINS
Amino acids:
Structure, Classification

Q.Give example of one suphur and –OH group containing amino acid.

Sulphur containing amino acid is Cysteine and –OH containing amino acid is Tyrosine.

General and Electro chemical properties of α-amino acids
Physiological importance of essential and non-essential amino acids
Proteins Bonds stabilizing protein structure
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Levels of organization

Q.What do you mean by α helicaltructure of a protein?

In 1951, Pauling proposed the α helix structure. The alpha helix (α-helix) is a common motif in the secondary structure of proteins and is a right hand-spiral conformation (i.e. helix) in which every backbone N−H group donates a hydrogen bond to the backbone C=O group of the amino acid located three or four residues earlier along the protein sequence. The alpha helix is also called a classic Pauling–Corey–Branson α-helix.

The name 3.613-helix is also used for this type of helix, denoting the average number of residues per helical turn, with 13 atoms being involved in the ring formed by the hydrogen bond (For further information follow contact zone).

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Protein metabolism:
Transamination

Q.What is transamination?

Transamination is a biochemical process of nitrogen catabolism which transfers the amino group of an amino acid (e.g., alanine) to a keto acid (e.g., α-ketoglutarate), changing the latter into a new amino acid (e.g., glutamate) and the original amino acid into a new keto acid (e.g., pyruvate).

General feature:

(1)Occurrence— Transaminase or aminotransferase catalyzes Transamination in mitochondria and cytoplasm of liver in particular and also of kidneys, hearts, testes and brain.

(2)Prosthetic group of the enzyme— Pyridoxal phosphate (PLP) acts as prosthetic group for transaminase enzyme.

(3)Restriction on enzyme activity— Mammalian transaminase acts specifically on L-amino acids, but not on their D-isomers.

(4)Limitations of transaminase activity—This enzyme activity is not applicable for some amino acids like, threonine, lysine, proline and hydroxyproline.

(5)Ping-pong type of substrate reaction— Transaminations are double-displacement type of substrate reactions. The two substrates, viz, an amino acid and an α-keto acid, bind separately and successively with the prosthetic group of the enzyme.

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Q.Give full form of SGOT and SGPT.

SGOT—Serum glutamate oxaloacetate transaminase

SGPT— Serum glutamate pyruvate transaminase

Deamination
Urea cycle
Fate of C-skeleton of Glucogenic and Ketogenic amino acids

Q.Define ketogenic and glucogenic amino acid. Give suitable examples.

A ketogenic amino acid is an amino acid that can be degraded directly into acetyl-CoA, which is the precursor of ketone bodies. In humans, two amino acids are exclusively ketogenic: Leucine and Lysine.

A glucogenic amino acid is an amino acid that can be converted into glucose through gluconeogenesis. In humans, the glucogenic amino acids are: Alanine, Arginine, Asparagine, Aspartic acid, Cysteine, Glutamic acid, Glutamine etc.

Q.Give example of few amino acids those are representing both ketogenic and glucogenic status.

Phenylalanine, Isoleucine, Threonine, Tryptophan, Tyrosine.

NUCLEIC ACIDS
Structure of Purines, Pyrimidines
Nucleosides and Nucleotides

Q.Distinguish between nucleotide and nucleoside.

A nucleoside consists of a nitrogenous base covalently attached to a sugar (ribose or deoxyribose) but without the phosphate group. When phosphate group of nucleotide is removed by hydrolysis, the structure remaining is nucleoside. Several nucleoside analogues are used as antiviral or anticancer agents.

A nucleotide consists of a nitrogenous base, a sugar (ribose or deoxyribose) and one to three phosphate groups. Malfunctioning nucleotides are one of the main causes of all cancers known of today.

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Nucleic Acid Metabolism
Catabolism of adenosine, guanosine, cytosine and thymine

Q. State the possible of catabolism of adenine nucleotide in liver.

In the liver and cardiac muscle, purine 5’-nucleotidase hydrolyzes adenylate to Pi and adenosine. Adenosine deaminase deaminates adenosine to inosine. This enzyme can also deaminate deoxyadenosine to deoxyinosine. Purine nucleoside phosphorylase next uses Pi to phosphorolyze inosine to ribose 1-phosphate and hypoxanthine. Deoxyinosine is similarly phosphorolyzed to hypoxanthine and deoxyribose 1-phosphate (For further information follow contact zone).

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ENZYMES
Nomenclature and classification

Q.Decode EC 3.4.11.4.

Every enzyme code consists of the letters "EC" followed by four numbers separated by periods. Those numbers represent a progressively finer classification of the enzyme. Preliminary EC numbers exist and have an 'n' as part of the fourth (serial) digit (e.g. EC 3.5.1.n3).

Tripeptide aminopeptidases have the code "EC 3.4.11.4", whose components indicate the following groups of enzymes:

EC 3 enzymes are hydrolases (enzymes that use water to break up some other molecule.

EC 3.4 are hydrolases that act on peptide bonds.

EC 3.4.11 are those hydrolases that cleave off the amino-terminal amino acid from a polypeptide.

EC 3.4.11.4 are those that cleave off the amino-terminal end from a tripeptide.

Cofactors
Specificity of enzyme action
Isozymes

Q.What are the isozyme?

Isozymes (also known as isoenzymes or more generally as multiple forms of enzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes usually display different kinetic parameters (e.g. different KM values), or different regulatory properties. The existence of isozymes permits the fine-tuning of metabolism to meet the particular needs of a given tissue or developmental stage. In biochemistry, isozymes (or isoenzymes) are isoforms (closely related variants) of enzymes.

Lactate dehydrogenase is a tetramer of 2 kinds of 35-kd subunits encoed by similar genes. These subunits are associated to form 5 types of tetramers (H4, H3M1, H2M2, H1M3, and M4. These species are called Isoenzyme or isozyme. [LDH1-in cardiac muscle, LDH2-in kidney, RBC etc].

Mechanism of enzyme action;
Enzyme kinetics

Q.What is turnover number?

The turnover number of an enzyme is the number of substrate molecules converted into product by an enzyme molecule in a unit time when the enzyme is fully saturated with substrate.

It is equal to the kinetic constant k3. The maximum rate Vmax reveals the turnover number of an enzyme if the concentration of active sites [Et] is known, because Vmax = k3 [Et].

A 106M solution of carbonic anhydrase catalyses the formation of 0.6M H2CO3 per second when it is fully saturated with substrate. Hence, k3 is 6×10-5s-1. Each round of catalysis occurs in a time equal to 1/k3 which is 1.7µs for carbonic anhydrase. The turnover number of most enzymes with their physiological substrates fall in the range from 1 to 104 per second.

Derivation of Michaelis-Menten equation
Lineweaver-Burk plot
Factors affecting rate of enzyme-catalyzed reactions; Enzyme inhibition.

Q.Mention the name of substrate and type of inhibitor for the following enzymes.

(a)Succinate dehydrogenase and (b)Citrate synthetase

Q.Mention the name of substrate and type of inhibitor for the following enzymes.

(a)Succinate dehydrogenase and (b)Citrate synthetase

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OXIDATIVE PHOSPHORYLATION
Redox systems
Mitochondrial respiratory chain

Q.Distinguish between substrate level phosphorylation and oxidative phosphorylation.

Substrate level phosphorylation-

Substrates are oxidizes via an NAD-linked dehydrogenase and the respiratory chain, approximately 3 mol of inorganic phosphates are incorporated into 3 mol of ATP per half mol of O2 consumed, i.e; the P:O = 3.

Oxidative phosphorylation-

Substrate is oxidized via a flavoprotein-linked dehydrogenase, only 2 mols of ATP are formed, i.e; P:O=2.

The main difference between substrate level phosphorylation and oxidative phosphorylation is that substrate level phosphorylation is a direct phosphorylation of ADP with a phosphate group by using the energy obtained from a coupled reaction whereas oxidative phosphorylation is the production of ATP from the oxidized NADH and FADH .

Substrate-level phosphorylation is directly phosphorylating ADP with a phosphate and energy provided from a coupled reaction. SLP will only occur if there is a reaction that releases sufficient energy to allow the direct phosphorylation of ADP. Oxidative phosphorylation is when ATP is generated from the oxidation of NADH and FADH2 and the subsequent transfer of electrons and pumping of protons. That process generates an electrochemical gradient, which is required to power the ATP synthase.

Inhibitors and un-couplers of Electron Transport System

Q.State inhibitory role of Rotenone and Atractylate in ETC.

Rotenone :It is the non-toxic inhibitors of Electron transport chain. These compound extracted from roots of tropical plant Derris elliptica and Lonchoncarpus nicou. It binds at Complex I between Fe-S protein and Ubiquinone.

This is non-toxic to mammals because poorly absorbed. Shows toxic effect in fishes.

Atractylate :It backs oxidative phosphorylation by compelling with ATP & ADP for a site on the ADP-ATP antiport of the mitochondrial membranes. One of the inhibitors list which blocks the oxidative phosphorylation.

 
 
 
 
 
 
 
 
 
 
 
 
 
 

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