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    BIOMOLECULES - BIOLOGY (Theory)

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    Ankith Murthy

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    Chapter Outline 9.2.1 Water Organic Compounds 9.3.1 Carbohydrates 9.3.2 Amino acids 9.3.3 Proteins 9.3.4 Lipids 9.3.5 Nucleic Acids Concept of Metabolism 9.4.1. Metabolic basis for living organisms 9.4.2 The living state 9.1 INTRODUCTION } G.N. RAMACHANDRAN, an outstanding figure in the field of protein structure, was the founder of the ‘Madras school’ of conformational analysis of biopolymers. His discovery of the triple helical structure of collagen published in Nature in 1954 and his analysis of the allowed conformations of proteins through the use of the ‘Ramachandran plot’ rank among the most outstanding contri- butions in structural biology. He was born on October 8, 1922, in a small town, not far from Cochin on the southwestern coast of India. His father was a professor of mathematics at a local college and thus had considerable influence in shaping Ramachandran’s interest in mathematics. After completing his school years, Ramachandran graduated in 1942 as the topranking student in the B.Sc. (Honors) Physics course of the University of Madras. He received a Ph.D. from Cambridge University in 1949. While at Cambridge, Ramachandran met Linus Pauling and was deeply influenced by his publications on models of the helix and B-sheet structures that directed his attention to solving the structure of collagen. He Passed away at the age of 78, on April 7, 200) How do Enzy high rat 9.9 Fact 9.10 Enzyme In 9.11 Cl Enz 9.12 Enzyme Cl Nomenclature 9.13 Co-factors me! sification Cell is the structural and functional unit of organisms. Cell performs a wide variety of biological activities. Cell consists of various cell organelles and each cell organelle performs variou biochemical activities. Cell components are ma up of some basic compounds which help in complex interactions. The various types of molecules that perform variety of functions put together called cellular pool. It consists of both inorganic and organic molecules. Inorganic component includes salts, mineral jons and water All the carbon compounds that we get from living organisms are called biomolecules. Organi component includes carbohydrates, proteins, lip! hormones, enzymes, vitamins etc, It is als important to study different types of elemen!’ Present in a living body. The composition 0! elements may vary from plants to animals (0! different organisms. We can compare this analys* with the analysis of elements present in the ea'th crust (non-living matter). Both may contain som similar elements in different percentages. In this chapter emphasis is given to study about variow macromolecules of the ¢ Table 9.1.1 A com; Parison of elem, present in non-livi ae NG and living matter 0.03 See od negligible study the various biomolecul Organisms, the tissues are grinded acetic acid (C/,CCOOH) in a mort fugation, the supernatent is called and residue is called acid insolu are further separated and les of living with trichloro- ‘ar. After centri- acid soluble pool ble pellet. These identified by various analytical techniques. Acid soluble pool contains low molecular weight molecules and they are called as biomicro-molecules. The acid insoluble pellet Contains high molecular weight molecules and they are called as biomacromolecules. The elemental analysis can be carried out by ash analysis. Dried biomass of organism is kept in silica crussibles These crussibles are placed in muffle furnace at 450°C. The ash thus obtained is analysed to find Sut the composition of elements with the help of atomic absorption spectrophotometer (AAS). Average composition of compounds in a cell Metabolites The living organism contains thousands of organic Compounds and these may be considered as metabolities. These are divided into primary and Secondary metabolites. Primary metabolites: These organic compounds are involved in basic metabolic activities eg. sugars, amino acids, ois, fats, nitrogen Secondary metabolites: Organic compounds other than primary metabolites are called secondary metabolites. Eg. alkaloids, flavonoides, rubber antibiotics, pigments etc, (Table 9.5.2). Their roles Or functions are not known clearly. Some secondary metabolites have ecological importance. However many of them are useful to human welfare, Table 9.1.2 Some secondary metabolites Pigments | Carotenoids, Anthocyanins, ete. Alkaloids | Morphine, Codeine, etc. Terpenoides Monoterpenes, Diterpenes ete Essential oils! Lemon grass oil, ete. Toxins + # | Abrin, Ricin Concanavalin A Vinblastin, curcumin, ete. Polymeric substances Rubber, gums, cellulose 9.2 INORGANIC SUBSTANCES ———SIIC'SUBSTANCES 0) Living cells contain 70-90% of water. In humans, about 2/3rd of body is formed by water. Water ic a tiny polar molecule. Fluidity of water is maintained by rapid formation and dissociation of H-bonds between water molecules, It acts as solvent in living organisms, Wai and is a source of Ht biochemical reactions. Di ler is a reagent & OH- for various uring photosynthesis, water is a proton donor, Due to solvent action ‘water helps to maintain constant pH. Due to the high specific heat and mobility of water, it distributes heat uniformly through out the body, High latent heat of vapourisation of water causes, excess heat and maintain constant ature, Elimination of waste products ne helps in homeostasis. Water is most hemical in Jiving organisms, | body temper | through urin { abundant cl 83. ORGANICCOMPOUNDS —_ Living organisms contain a wide variety of organic compounds. Some of them are described below. 9.3.1 Carbohydrates These are polyhydroxy aldehydes (or) ketones They are also known as saccharides as they are formed of sugars. They are divided into 1) Monosaccharides 2) Oligosaccharides 3) Polysaccharides 1, Monosaccharides They consists of a single polyhydroxy aldehyde (or) ketone acid, Generally they are sweet in taste and soluble in water. The emperical formula is (CH,O)n, A monosaccharide with aldehyde group is called aldose and with ketone group is called ketose. They are further divided into different groups based on the number of carbon atoms. The most abundant monosaccharide in nature is D-glucose. (Fig. 9.3.1) ‘CH,OH al c-0 3 HO —=C—H | ®CH.OH H Dihydroxyacetone Fig. 9.3.1 Diagrams showing different monosaccharides i) Trioso: ‘These are smallest monosacchariy., & contain three carbon atoms. These a), intermediates of respiration, Ey. Dil, acete 1¢ (Ketose) Glyceraldehyde (Aldos. {i) Tetro: They have four carbon atom, These are intermediates of photosynthesis respiratory pathways. Eg. Erythros. Threose (aldose), Erythrulose (ketose lil) Pentoses: They have five carbon atoms pe, molecule, Eg, Ribose, Deoxyribose, Xylulose, Arabino, (a jose) Ribulose (ketolose). iv) Hexoses: They have six carbon atoms pe; molecule, Eg. Glucose, Galactose, Mannose (aldos Fructose (Ketose) Glucose is called dextorse or corn sugar (or) grape sugar which is a common respiratory substrate, Fructose is common fruit sugar or levulose and is sweetest of all natural sugars with an index of 170. Galactose is a constitutent of agar-agar, Manitol is an alcohol of mannose found in brown algae. v) Heptoses: They have seven carbon atoms pe: molecule, Eg. Sedoheptulose (ketose), an intermediate of photosynthetic pathway. 2. Oligosaccharides These are small sized polymers of monosac charides having 2-9 simple sugar units. They are classified into different groups based on the number of units. (Fig. 9.3.2). A) Disaccharides These are smallest oligosaccharides formed by condensation of two monosaccharides. (Fix. 9. eg, Maltose = Glucose + Glucose (reducing sug!) Lactose = Glucose + Galactose (reducing sugat) Sucrose = D-Glucose + D-fructose (non-reducing sugar) Trehalose = Glucose + Glucose Cellobiose = Glucose + Glucose On HH CHOH eae CH.OH Lactose ‘Oo. ‘OOH (Galactose-glucose) H oH u OH CH:OH CHOW Maltose ° 0. OH (Glucose-glucose) Ht HS OH OH Fig. 9.3.2 i) Maltose: It is known as malt - sugar. Two glucose molecules are held by a(1—> 4) glycosidic bond. It is formed of starch. It is found in germinating cereals, ii) Sucrose: It is also called as cane sugar or table sugar. It is obtained from stem of sugarcane and roots of sugar beet. It does not Possess free aldehyde (or) ketone group and hence it is a non-reducing sugar. ili) Lactose: It is also known as milk sugar and abundantly found in milk. Glucose and galactose are linked by B(1 4) linkage Souring of milk occurs during fermentation, where lactose is converted into lactic acid by Lactobacillus bacteria. B) Trisaccharides They are made up of three sugar units. Eg. Raffinose (=Melitose) = & Galactose +OGlucose, + B fructose C) Tetrasaccharides They are made up of four sugar units. Eg. Stachyose = Galactose, +0 Galactose. +0Glucose, +8 Fructose D) Pentasaccharides They are made up of five sugar units. IEg. Verbascose = Galactose, + 01 Galactose, +0 Galactose, +0 Glucose +f Fructose Reducing sugars: Sugars having free aldehyde or keto groups are called reducing Sugars. The are able to reduce cupric ions Cut? cuprous ions Cut. Benedict's test and Fehling’s test The solution is blue in colour, after adding sugar to the solution it is heated mildly. Orange to brick Ted precipitate of cuprous oxide (Cu*) is observed. Eg. All monosaccharides, disaccharides like lactose and maltose. Non-reducing sugars : They do not have free aldehyde or keto group. They cannot change cupric ions to cuprous ions. Eg. Sucrose, Stachyose. 3. Polysaccharides These are long chain carbohydrates with large number of units. They are also called glycons and Not sweet to taste. Structurally they are two types viz 1) Homopolysaccharides 2) Heteropolysaccharides 1) Homopolysaccharides These are complex carbohydrates that are formed by polymerisation of only one type of mono- saccharides. Eg. Pentosan - Polymer of pentose Hexosan - Polymer of hexose Araban — - Polymer of arabinose Xylan Polymer of xylose Similarly Glucosan, fructosan, starch, glycogen, cellulose. 2) Heteropolysaccharides These are carbohydrates formed by condensation of more than one type of monosaccharide monomers. Eg. Hemicellulose, Pectin & Muco- polysaccharides, Polysaccharides are functionally three types. viz. storage, structural and mucopoly saccharides. 1) They function as reserve hy and biosynthesis. Eg. Starch, i) iii) Inui Storage polysaccharides food which can be quired for respiration Glycogen, Inulin. (Amylum = starch) It is a glucosan homopolysaccharide. It is the major reserve food material in plants. Starch is the end product of photosynthesis and stored in the form of microscopic grains in amyloplasts. Starch has two components. Starch forms helical secon- dary unlike cellulose structures holding 1, molecules in it. The starch iodine is blue in colour, drolysed to form sugars rei Starch: Glycogen: It is also known as animal starch. It ts the resreve food material in animals, bacteria and fungi. It is formed from glucose through glycogenesis. Glycogen has a(1 > 4) linkages in the straight part and a(1 36) linkages in the branches. It gives red colour wiht iodine solution it is the smallest polysaccharide made up of fructose. It is formed by B (2,1) glycosidic linkages. It is the reserve food material found in Asteraceae members. 2) Structural polysaccharides i) Cellulose: It is a fibrous glucosan homopoly- saccharide and a structural component of cell wall in all plants and some fungi. It forms 50% ‘of organic carbon of plants. It is also the most abundant organic molecule of earth. Cellulose is a linear, unbranched straight molecule in which adjacent f- glucose units are joined by BC — 4) linkages. It does not react with iodine due to the absence of helices. It gives purple colour with zine chloride-iodine. Importance 1. Cellulose rich wood is used in production of paper. 2. Cellulose is used in textiles, preparation of bags, ropes. 3. Cellulose nitrate (nitro cellulose) is used in propellent explosives. . Carboxy methyl cellulose is used as emulsifier and smoothening reagent of ice creams, medicines and cosmetics. 5. Rayon and cellophane are made up of cellu- lose xanthate. ii) Hemicellulose: It is a heteropolysaccharide D-xylose, mannose, glucose, galactose. Hey cellulose consists of either pentoses or hexosc, They form link between pectic compounds «a, cellulose microfibrils ill) Pectin: It is a heteropolysaccharide and mad up of arabinose, galactose and galactouron acid. It is water soluble and occurs in middl: lamella and cell wall of plants iv) Chitin: It is a structural homopolysaccharide It is found in fungal wall and exoskeleton arthropods. Chitin is insoluble in water. It is polymer of N-acety glucosamine and mon, mers are joined by f(1— 4) linkages. 3) Mucopolysaccharides They are heteropolysaccharides of hic: molecular weight. They are formed fron galactose, mannose, sugar derivatives and uroni acid. They are found inside the plant cell wall, outside the cells or bodies of bacteria, bluc green algae and many aquatic plants. a) Agar-Agar is a cell wall mucopoly saccharide of some red algae like Gelidium Gelidiella, Gracilaria. Agar is used as culws medium, thickner, stabilizer and laxative b) Carrageenin (from Chondrus crispus (10 moss)) used as emulsifer and cleaning agen ©) Heparin is a highly sulphated mucopol) saccharide made up of alternate units « glucosamine and glucuronic acid. It \ abundant in blood and connective tissue. | is used as blood anti-coagulant 9.3.2 Amino acids Amino acids are organic compounds conti" an amino group and an acidic group as substi ‘on the same carbon i.e., the o-amino acids. 1! are substituted methanes, There are four subs!" groups occupying the four valency positions. T are hydrogen, carboxyl group, amino grouP variabloe group designated as R group. 5: the nature of R group there are many 20° wef, those Which occur in proteins are only twenty types: the R group in these proteinaceous no acids could be a hydrogen (the amino acid j called glycine), @ methyl group (alanine), hydroxy methyl (Serine), ete., Three of the twenty ye shown in Fig. 9.3.3. COOH rook COOH | ! H-C—NH, Bias Nie H—C—NH, | I H Cs Glycine Alanine Serine Fig. 9.3.3 Amino acids ‘he chemical and physical properties of amino acids are essentially of the amino, carboxyl and the R functional groups. Based on number of amino and carboxyl groups, there are acidic (e.g., glutamic acid), basic (lysine) and neutral (valine) amino acids. Similarly, there are aromatic amino acids (yrosine, phenylalanine, tryptophan). A particular of amino acids is the ionizable natured of -NH, and -COOH groups. Hence in solutions of different pH values, the structures of amino acids will change. Amino acids are aminated (-NH,) mono or dicarboxylic acids. Their general formula is R-CH-NH,-COOH. They are made up of C, H, 0, N and some of them contain “S” also. There are nearly 700 types of amino acids out of which 20 types of amino acids occuring in proteins. They ae highly polar. In aqueous solution, the carboxyl group transfers a proton to NH, group to give Positive and negative charges within the molecule. This type of species is called Zwitter ion or inner salt, R R , 1 H,N~CH-COOH = NH} ~ CH-COO™ 2witter ion In acidic solution, amino acid becomes positive ion and in basic solution negative ion. Amino acids are divided into eight types based m their structure and reaction (Table. 9.3.3) 1) Acidic amino acids: They have non cyclic hydrocarbon chain with one amino group and two carboxylic groups. Eg. Glutamic acid, Aspartic acid. 2) Basic amino acids: These amino acids have non cyclic hydrocarbon chain with two amino groups and one carboxylic groups. Eg. Lysine, Arginine The acidic or basic nature is due to presence of either amino group or carboxylic group on the side chain, 3) Neutral amino acids: They have non cyclic hydrocarbon chain, with one amino group and one carboxylic group. Eg. Glycine, Alanine, Valine, Leucine 4) Sulphur containing amino acids: They have sulphur in their structure Eg. Methionine, Cysteine 5) Alcoholic amino acids: An alcohilic or hydroxyl group is present in the amino acids. Eg. Serine, Threonine 6) Aromatic amino acids: A cyclic or ring structure is found in hydrocarbon side chain, Eg. Tyrosine, Phenylalanine, Tryptophan. 7) Heterocyclic amino acids: A hetrocyclic ring structure occur in the hydrocarbon side chain of the amino acid, Eg. Tryptophan, Histidine 8) Imino acids: There are amino acids in which (NH) is present instead of amino group. Eg. Proline 9.3.3 Proteins The term protein was coined by Berzelius. The word protein is derived from Greek word “Proteios” which means prime importance. Proteins are naturally occuring polypeptides that contain more than 50 amino acid units and generally 100-300 units therefore, a protein is a heteropolymer, Eg. Silk, hair, connective tissues, most of the enzymes. CHa (CH-CHNH~Co 9}, Valine Val y CHy 4 [ HC. cece, CHNH,~co0, Leucine Leu i HC GHs. ~ ‘CH- CHNH,~ Isoleucine lle I CH, ~ 2- COOK Aspartic acid ‘Asp D HOOC - CH, ~ CHNH, ~ C00); Asparagine(Amide) ‘Asn N HOOC - CH, - CHNH, ~ Coy}, Glutamic Acid . Glu E HOOC ~ CH, ~ CH, =CHNH « | Glutamine(Amide) Gin Q BOGS CH, = CH RQ Gas ns K H,NCH, - (CH,), ~ CHNH, — Co, I, Arginine Arg R H,N-C-NH-(CH)),-CHNH,-C00} Cysteine Cys Cc HS - CH, - CHNH, ~ COOH Methionine Met M CH, - S~ CH, - CH, ~ CHNH Serine Ser s HOCH, ~ CHNH, ~ COOH Threonine Thr a CH, - CHOH ~ CHNH, ~ C0011 phen yiaiation Phe F O~ arcs, ‘00H Tyrosine Tyr Y wo{\- (CH,-CHNH,-COO! Histidi cy fear (CHp-CHNH,-COOH HN N Functions Intercellular ground sub: ‘| Enzyme mee Hormone agents Sensory reception (smell, taste, hormone, etc) Enables glucose transport into cells na polypeptide chain “N” terminal is written on left side and ‘C’ terminal on the right hand side to the chain. Collagen is the most abundant protein of the animal kingdom and RuBisCO is the most abundant protein in biosphere. 21% and 22™ Amino acids ‘Selenocysteine (21% selenium — cysteine) Pyrrolysine (224) 9.3.3.1 Structure of protein Proteins, as mentioned earlier, are heteropolymers containing strings of amino acids. Structure of molcules means different things in different contexts, Jn organic chemistry, the structure invariably refers to the molecular formulae (Eg. NaCl, MeCl., etc) Organic chemists always write a two dimentional view of the molecuels (Eg. benzene, napthalene. eie,). Physicists conjure up the three dimentional views of molecular while biologists describe the protein structur at four levels. (Fig. 9.3.4) Alanine Asparagine Aspartate Cysteine Glutamate Glutamine Glycine Proline Serine Tyrosine Alpha-Helix Beta-Plated sheet (c) Tertiary Hydrogen Disultide bond Quaternary Fig. 9.3.4 1. Primary structure Linear polypeptide chain which is linked by peptide bonds gives primary structure of protein. Any change in the sequence of an amino acid produces a different protein. In the primary structure, first amino acid is “N” terminal and last amino acid is “C” terminal. The distance between two adjacent peptide bonds is about 0.3Snm. Primary structure does not give any information about the functional properties of protein. (Fig. 9.3.5). Amino end Fig. 9.3.5 Primary structure of protein 2. Secondary structure It is developed from primary structure by their peptide bonds through formation of intra or inter Polypeptide hydrogen bonds, which make proteins functional. (Fig. 9.3.6) Secondary structure is of three types. __ ee (3) 0 -Helix; (ii) B -pleated shee ; (iii) Cottagen helix (a) o S .6 A Secondary Structure i) G@-Helix : A polypeptide chain which forms a regular spiral or coil around an imaginary axis is called oc-helix. Some of the structures which show GL-helix are keratin of hair, fur, claws, hooves and feathers. The coils of or-helix are held together by hydrogen bonds between amino acids of sucessive truns of the coil parallel to the main axis of the fibre. “R” groups occur towards the outer side of. Gt-helix. The ot-helix has a pitch of 5.4A°. Each tum has 3.6 amino acids. ¢¢-helix is found in both fibrous and globular proteins. (Fig. 9.3.7(a)), ii) B pleated sheet : When two or more peptide chains are inter connected by hydrogen bonds to produce a pleated sheet like structure known as pleated sheet. Chains may be parallel or antiparallel, Straight hydrogen bonds occur between imide group (-NH) of one poly peptide and carboxyl (-CO-) group of adjacent polypeptide. Cross linkages help in stabilisation of pleated sheet. In parallel chain “'N” atoms of adjeent strands of poly-peptides point in the same directions (B Keratin of feathers). In antiparallel B pleated sheet, “N” atoms are in opposite directions (silk, fibroin) (Fig. 9.3.7 (byc)) -Helix Fig. 9.3.7 (a) Parallel 6 ~ Conformation nen, * s HCR N A REY Soret Y= 0-- mes, NN Hin RU ice on ae ~ noe Z ‘ ze Antiparaliel B~ Conformatios Fig. 9.3.7(b) NY) B pleated sheet Fig. 9.3.7(c) Secondary structure of protein lil) Collagen helix: Collagen has large amount of glycine and proline hence it cannot form. t-helix. Three polypetide chains come together and Hydrogen bond ionic bond ae i (A) Fig. 9.3.9(A) Tertiary-structure Tun parallel to form a ri 'S stabilized by hydrog. The triple helix of collay tis present in connecti Fig, 9.3.8) ight handed super helix that jen bonds among the three gen is called tropo collagen Ve tissue tendons and bones Fig. 9.3.8 Structure of collagen G.N. RAMACHANDRAN was an outstanding figure in the field of protein structure. He discovered the triple helical structure of collagen. 3.Teritary structure Ivis the three dimensional arrangemnt of all atoms in the protein. In this structure polar side chains are exposed and non polar amino acid side chains are hidden inside the coils and folds. Teritary structure is maintained and stabilized by non covalent weak bonds like hydrogen bonds, ionic bonds, hydrophilic and hydrophobic bonds, disulphide bonds, vander waals interactions. Fig. 9.3.9 & 9.3.9), Eg. Myoglobin a -chain B-cha (B) (B) Quaternary Structure 4. Quaternary structure Some proteins are an assembly of more than eee polypeptide or subunits, The manner in which individual folded polypeptides or subunits, are arranged with respect to each other (¢.g. linear string of spheres spheres arranged one upon each other in the form of a cube or plate etc) is the architecture of a protein otherwise called the quarternary structure of a protein, Adult human haemoglobin consists of 4 subunits. Two of these are identical to cach other. Hence, two subunits of a type and two subunits of B type together constitute the human haemoglobin (Hb). These involve two or more identical or non identical polypeptide chains linked by weak non covalent bonds. (Fig. 9.3.9B), Eg, Haemoglobin Fig. 9.3.9 Bonds in 1°, 2°, 3°, 4° protein (bonds | Peptide bonds | H-bonds, hydrophobic bonds Van der wall interaction. H-bonds, hydrophobic bonds | ‘Van der wall interaction, Ionic = ___| bonds, disulfide bonds. 4 H-bonds, hydrophobic bonds Van der wall interaction, Ionic bonds, disulfide bonds. ee i | (8OT4y 9.3.4 Lipids (Gk lipos - Fat) mT leide: are Cone orne We Doe lipids are not strictly ,, molecular ounds an fs ee They are soluble in non-polar «o, because they contain relatively long hydro... chains that are non-polar and thus hydroph., shaken with water they often form small or ‘micelles’ and it is a modified colloidal co, called emulsion. Lipids are generally water insoluble. 1), could be simple fatty acids. A fatty acid ), carboxyl group attached to an R group. The R yi... could be a methyl (-CH,), or ethyl (-C,, higher number of -CH, groups (I carbon i |, carbons). For example, palmitic acid has 16 carly including carboxyl carbon. Arachidonic acid } 20 carbon atoms including the carboxy! cary Fatty acids could be saturated (without doutic bond) or unsaturated (with one or more C=C dovhi: bonds). Another simple lipid is glycerol which tihydroxy propane. Many lipids have both glyc and fatty acids. Here the fatty acids are foun: esterified with glycerol. They can be the: | monoglycerides, diglycerides and triglycerides These are also called fats and oils based on meltin: point. Oils have lower melting point (ly. ©: oul) and hence remain as oil in winters. Some lipis have phosphorous and a phosphorylated organi: compound in them. These are phospholipids. The) are found in cell membrane. Lecithin is on example. Some tissues especially the neural tissues have lipids with more complex structures (Fig. 9.3.10). 9.3.4.1 Fatty Acids Organic acids having single carboxylic group ané 1-19 carbon atoms are called fatty acids. Common fatty acids have even number of carbon atoms Carboxylic group is polar due to CO and OH an! it is hydrophilic, ‘The remaining part of fatty acid is hydrophobi: and non polar. They are amphipathetic as it has are further divided into two types CH3—(CH2)14—COOH, ° z CH, Fatty acid 20H tl (Palmitic acia) Hee, a i GSO Rcea | '2—C—O—CH ° CH.—o | Il G 4 CH,—O—C—R, & lycerol Tiigvosrael are if zmO—C—A, and Rg are fatty acids) 2 i ° I iO OCH, cH, | ya EX Phospholipid (Lecithin) 3 cH Gy a Fig. 9.3.10 Fats 4) Saturated fatty acids In these fatty acids all the carbon atoms of the hydrocarbon chain are linked by single bonds and the general formula is C,H,,0,. Bg. Palmitic acid = - CH,(CH,), ,COOH.(16C) Stearic Acid CH,(CH,), ,COOH.(18C) Arachidic Acid - CH,(CH,),,COOH.(20C) 2) Unsaturated fatty acids These fatty acids contain one or more double bonds. General formula is C,H,, »,0, (x is number of double bonds). The melting point of unsaturated fatty acids decreases with increase in umber of double bonds. Saturated fatty acids are solid even in summer. Unsaturated fatty acids are fatty converted to saturated acids by acids which cannot be synthesized in called essential fatty acids. acid, Linolenic acid, Arachidonic fatty acids which can be synthesized in non-essential fatty acids. acid, Stearic acid Cholesterol and Oils (lipids) Polyunsaturated fatty acids Fatty acids with more double bonds are called Polyunsaturated fatty acids (PUFA)..These are |. very useful for human beings in retarding Athero- sclerosis, leads to reduction of cholesterol in blood. So these are very useful for patients suffering from high blood cholesterol, hyper- tension ete. Eg. (1) Oleic acid (18C), (2) Linoleic acid (18C), 3) Linolenic acid (18C) (4) Arachidonic acid 20C), Sunflower oil and Safflower oil contain more polyunsaturated fatty acids. 9.3.4.2 Types of Lipids 4) Simple lipids These are esters of fatty acids with different alcohols. Fats or Oils: These are triesters of fatty acids and glycerol (trihydroxy propane). They are called triglycerides because glycerol forms ester linkages with three fatty acid molecules. ° i CH,OH +R\COOH — CH,»—O—C—R, I CHOH +R,COOH —= I CH,OH + RyCOOH CH.—O—C—Ry ‘Triglyceride is oF conjugated lipids th some other substances: compounds having Pid and phosphoric acid. Ovt of groups of glycerol (wo are attached © to phosphoric acid. led membrane lipids as in cell membrances. 2) Compound Lipid These are the lipids wi Phospholipids: They are glycerol, fatty three hydroxyl to fatty acids and on Phospholipids are also cal they are present as bilayer i (Phosphatidy! ethanolamine): nsulating material Eg.a) Cephalin Found in brain and acts as 1 for nerves. thin (Phosphatidyl choline): They are (b) Lecit permeability and involved in regulating cell n of cells. (Fig. 9.3.10). osmotic tension cH, — OOC-R: -H ° eaecall 4, -0— P—O CHa CHa NICH)s Glycerol + Fatty acid * Fig. 9.3.11 Structure of Lecithin Steroids (Gk: Stereos = Solid) tty acids, hence they are non saponifiable. All steroids are derivatives of a fused ‘and fully saturated ring system called “Sterane”. This system consists of 3 cyclohexane rings (A, B. Jinear manner and a terminal They donot contain fa C) fused in nonl cyclopentane ring (D). Cholesterol (C,,H,gOH): It is fat soluble steroid alcohgl. It has 17 carbon nucleus of four fused hydrcarbon rings. Hydroxyl group is found on 3rd carbon, two methyl groups at carbon position 10 and 13. The side chain consists of 8 carbons and is attached at carbon 17. Its parent hydrocarbon is Cholestane (CyyH,,). Cholesterol is absent in vegetable oils. It is found in saturated fats of animals origin (Eg. butter, ghee and egg yolk), but vamaspati does not contain cholestrol. Bulk of it is synthesized in the body. Unsaturated * blood cholestrol levels ce synthesis. Cholesterol is digest Cholesterol esterase Present, «, Normal level of cholestr! i eryod is 140-250 me/100 Tl ‘There are tWo types of cholestrol (Fig. 9.4.12). Fig. 9.3.12 Structure of Cholesterol Nature of bond linking monomers in a polymer In a polypeptide or a protein, amino acids are linkes by a peptide bond which is formed when the carboxyl (COOH) group of one amino acid reacts with the amino (-NH,) group of the next amino acid with the elimination of a water moiety (the process is called dehydration). In a polysaccharide the individual monosaccharides are linked by ® glycosidic bond. This bond is also formed by dehydration. This bond is formed between (wo carbon atoms of two adjacent monosaccharides In a nucleic acid a phosphate moiety links the 3 carbon of one sugar of one nucleotide to the 5” carbon of the sugar of the succeeding nucleotide The bond between the phosphate and hydroxy! group of sugar is an ester bond. As there is one such ester bond on either side, it is called phosphodiester bond. 935 Nucleic acids: Jeic acids are Macromolecules co, ae O, N and P. Nucleic acids were fin isis oy sss pysician Freldrich Miescher fom ne! Repeat sed nucleo, Richard Alsat renamed muclein as nucleic acid, Kosse) wn Hdentified two types of nucleic aces pro and RNA. Usually both of them occur in a np Vises are the Only entities which have only snc typeof nvcleic acid i.e, either DNA or Rove 9.3.5.1 Chemical composition Atbert Kossel identified the constituents acids as nitrogen bases, 5 - phosphoric acid. (Fig. 9.3./3) 4) Phosphoric Acid (H,PO,) Phosphoric aa has three reactive hydroxyl groups (OH) of which two are involved in forming sugar. back bone. The phosphate moiety binds to the 5' carbon of one pentose sugar and 3icarbon of neighbouring pentose molecule to produce phosphodiester linkage. Phosphate moeity carries negative charge. of nucleic carbon sugar and Hh Pyrimidine bases in RNA & gee u 5 Purine bases HOH,C HM HOH,C r 0. 0. ‘Ne m7| 1 4 wi Hu 4 *on 7 On on oH m_ showing structu Fig, 9.3.13 Diagra ane 2) Pentose Sugar Wo types of Sugars are present in nucleic acids namely ‘®) Ribose (C,H,,0,) present in RNA b) Deoxy ribose (C,H,,0,) present in DNA. Sugar molecules contain furanose ring structure. The oxygen atom present at the second carbon of ribose is absent in deoxyribose giving NS name 2' deoxyribose. Deoxyribose is nonreactive and stable than ribose molecule (Fig. 9.3.13) 3) Nitrogen bases The organiv bases present in nucleic acids are heterocyclic compounds containing nitrogen in their rings. Nitrogen bases are of two types namely Purines and pyrimidines (Fig. 9.3.13). i) Purines: Purines are nine - membered dicyclic or double ring and heterocyclic nitrogen bases where the two rings are joined at 4 & 5 positions, Nitrogen occurs at 1, 3, 7 and 9 positions, Purines are of two types, adenine (A) and guanine (G). 5 banter * s of Purines, Pyrimidines, ribose and eee vse 80TH, ii) Pyrimidines: They are 6-membered, mono- cyclic or single ring and heterocyclic nitrogen bases. They have nitrogen at 1 and 3 positions. Pyrimidines are of three types-Cytosine (C), Thymine (T) and Uracil (U). DNA contains : A, G, T and C RNA contains : A, G, U and C Nomenclature and molecular formulae - CAN. Guanine - 60xy2-amino purine - C5H,N,O Thymine- 2,4 dioxy 5 methyl pyrimidine (S-methyl uracil) ~ CJH,N,O, Cytosine-2 oxy 4 amino pyrimidine-C,H,N,O Uracil - 2,4dioxypyrimidine - C\H,N,O, Nucleoside: An association of pentose sugar and a nitrogen base is called nucleoside. Nitrogen ‘base and pentose sugar are held by a glycosidic bond (—C-N-C) formed between carbon atom V' of pentose sugar with 1! Nitrogen atom of Pyrimidine or 9! nitrogen atom of purine. Depending upon the pentose sugar, nucleosides are of two types namely ribose nucleosides (4 types) and deoxy ribose nucleo-sides (4 types). Ribose + Adenine = Adenosine - A Ribose + Guanine = Guanosine - G Ribose + Cytosine = Cytidine - C Ribose + Uracil = Uridine - U Deoxyribose + Adenine = deoxyadenosine - dA Deoxyribose + Guanine = deoxyguanosine dG Deoxyribose + Cytosine = deoxycytidine - dC Deoxyribose + Thymine = deoxythymidine - dT Nucleotide: Phosphorylated nucleosides are called nucleotides. They are formed by association of pentose sugar, nitrogen base and phosphoric acid. Ribonucleotides: Adenine - 6amino purine Adenosine monophosphate AMP (Adenylic acid) Guanosine monophosphate GMP (Guanylic acid) Cytidine monophosphate CMP (Cytidylic acid) Uridine monophosphate UMP (Uridylic acid) Deoxyribonucleotides: Deoxy adenosine monophosphate dAMp Deoxyguanosine monophosphate dGMp Deoxythymidine monophosphate dTMp Deoxycytidine monophosphate dCMP 9.3.5.2 Deoxy ribonucleic acid (DNA) DNA is the common nucleic acid present in prokaryotic and eukaryotic cells. It-is not see; some viruses. i.e., RNA viruses. Avery, Maclevy and Me Carty presented first evidence that Dy, is genetic material by genetic transforma; experiment, Hershy and Chase showed that ¢) DNA is the carrier of genetic information fr, parent to offspring. Hence DNA is also call: “Chemical basis of heredity” or “Master chemic,) of cell” or “store house of genetic information Occurence In eukaryotes DNA is mostly found in nucleus constituent part of chromosome and in nucleolus (nuclear DNA). Small quantities of DNA is also sec in mitochondria and plastids (organelle DNA Nulcear DNA is associated with histone proteins to from chromatin fibres. Chromosomes arc condensed chromatin fibres. Organelle DNA | without histone proteins and is called naked DN.\ In prokaryotes DNA is mostly located in nucleoii and small quantities in cytoplasm as plasmids Viruses have either DNA or RNA but never both Most of the plant viruses have RNA as genet material and most of animal viruses possess DN.\ .Length of DNA The length of DNA is usually defined as numbe of nucleotides present in it. It is the characteris! of organism and its chromosomes. Bacteriophage $X174 (ss DNA) - 5386 nucleotides Bacteriophage lambda - 48502 base pairs Escherichia coli - 4,6x10° base pairs Haploid human cell ~ 3.3x10” base pairs Structure of DNA Watson & Crick elucidated the double helix mode! of DNA based on the earlier investigations mae by Erwin Chargaff, Franklin and Wilkins >) img X-ray diffraction techingue. a., to double helix model, DNA is composes cae v0 complementary polynucleotide strands wih are gal eile around one another in righ neyo ee These ‘wo strands are antiparlel to earn zt means if one chain has polarity Soa the oer has 3'-5\ In each back bone car” the nucleotides are joined by phosphodieac, ester bond is formed between 3.1 of a sugar molecule and Sth carbon of nex He. Both strands are held togey bonds between one purine in one strand a pyrimidine on the opposite strand. The je of these hydrogen bonds was firs: observed by L.C. Pauling. Adenine is bounded to thymine by two hydrogen bonds (A=T), Guanine is bounded to cytosine by three hydrogen bonts (G#O).As a result always a purine comes oppovite toa pyrimidine. This generates approximately uniform distance between the two strands of the elix, Thus DNA double helix resembles a twisted carbon sugar her by 5 =< major groove Fig. 9.3.14 DNA structure ladder. The back bone is formed by alternately arranged sugar and Phosphate and the steps are formed by complimentary base pairs (Fig. 9.3.14) Chargafr rule: E.E. Chargaff worked on chemical components of DNA and put forward Certain 8eneralisations, 1) Purines and Pyrimidines occur in equal amounts, 2) Purines of DNA Pyrimidines of DN. 3) A+G=T eG) are adenine and guanine. ‘A are thymine and cytosine. 4) Deoxyribose sugar and phosphate residues occur in equal number. 5) Purine adenine is equimolar with pyrimidine thymine 6) Purine guanine is equimolar with pyrimidine cytosine A+T 7) Base ratio CG is specific for a species. It does not change with age or tissue. The ratio can be used to identity the species. Eg. Human fiver iF) E. coli 0.93 T bacteriophage = 1.86 Dimensions of DNA (B-DNA) * The diameter of DNA helix is 20A°or 2nm. * The length of DNA depends upon the number of nucleotides, % The angle of each spiral is 360°. * Each helix includes one minor and one major groove, ach turn contains 10 bp or 20 nucleotides. * The angle separating the two successive nucleotide pairs is 36° * The pitch or length of each turn is 344° * The distance between the two successive base pairs in helix is 0.34 nm or 3.4A°. Types of DNA 4) Based on number of polynucleotide strands, DNA is of two types - ss DNA (single stranded DNA) and ds DNA (double stranded DNA) 8s DNA : It is seen in some viruses such as $%174 Bacteriophage and M-13 bacteriophage. ds DNA: It is common type of DNA seen in all eukaryotic and prokaryotic cells and also in some viruses such as T even bacteriophages. b) Based on type of helix DNA is of five types. A, B,C,D& Z Linear and circular DNA: linear DNA is one in which the ends are free. Linear DNA occurs in eukaryotic nuclei. In prokaryotes, chloroplast and mitochondria the DNA is circular, Properties of DNA 1,DNA has very high molecular wei, several million Daltons, 2. DNA has:special ability to absorb {5 a wave length of 260 nm 3, When exposed to high tempe: the two strands of DNA sepa: other. This phenomenon is called den, tion or melting, It is caused by hydrogen bonds between the ni Of the two strands, DNA areas get denatured easily and ar melting areus. Regions rich in ¢ high melting areas. This is because has only two hydrogen bonds where ay ; pair has three hydrogen bonds. The tv, of DNA separated by denatura: tendency to reassociate is called renatur,, = . It is a hydrophilic molecule and through cell membrane. 9.3.5.3 Ribonucleic acid (RNA) Occurence Both prokaryotic and eukaryotic cells besides DNA. How ever viruses cont of them. Most of the plant viruses h: most of the animal viruses have DNA RNA Viruses TMY, Tulip yellow mosaic viruses, wound ‘x viruses, influenza viruses, polio viruses ex In cukaryotes, RNA is chiefly con ribosomes. Traces of RNA are found in & mitochondria (semi autonomous cell o It is also seen in nucleus of eukaryote karyotes it is seen both in nucleiod and cy: Structure RNA generally consists of a single poly- strand. Each nucleotide contains three compo: namely phosphate group, ribose sugar (C.! and nitrogen bases. There are four types of = bases in RNA namely adenine, guanine, cytosine. Hence four types of nucleosides and 'ypes of nucleotides are present in RNA Present in RNA in place of thymine of DNA differs from thymine in lacking a methy! 2 (BOTANY) (CH,) at 5°C position. RNA generally being single ded, doesnot exhibit complementarity. Hence purines and pyrimidines donot exist in 1:1 ratio types of RNA RNA can be broadly classified into the types following RNA Non genetic RNA dsRNA mRNA rRNA TRINA. snRNA Some viruses do not contain DNA but only RNA. In them RNA is the sole genetic material and is called genetic RNA. Genet RNA is of two types, ss RNA and ds RNA ss RNA: RNA of viruses is generally single stranded. It does not exhibit complementarity, hence chargafi’s rule is not applicable, ss RNA my be straight or form pseudohelix. eg. TMV. ds RNA: In some viruses RNA is double stranded. The two strands are complimentary to each other and form a double helix as in ds DNA. Purine and pyrimidines exist in 1:1 ratio. Eg, Wound tumour virus, reo virus, rice dwarf virus, maize rough dwarf virus, blue tongue virus, bacteriophage 96, Rota virus Non genetic RNA: Cells which possess DNA as genetic material contain another type of RNA called non genetic RNA. These RNAs depend on DNA for their synthesis (transcription) and are involved in protein synthesis. Non genetic RNA is, of four types, m RNA, r RNA, t RNA and SnRNA. ___ 9.4. CONCE! SEPT OF METABOLISM A simple bacterialcell, a protozoan, a plant or an animal cell, contain thousands of organic compounds. These compounds or biomolecules are Present in certain concentrations (expressed, as mol/ Cell or mols/litre etc.). One of the greatest discoveries ever made was the observation that all these biomolecules have a turn over. This means that they are constantly being changed into some other biomolecules and also made from some other biomolecules. This breaking and making is through Shemical reactions constantly occuring in living tic organisms. Together all these chemical reactions are called metabolism. Each of the metabolic in the transformation of biomolecules, A few examples for such metabolic transformations are: removal of CO, from amino acids making an amino acid into an amine, removal of amino group in a nucleotide base; hydrolysis of a glycosidic bond in a disaccharide; etc. We can list tens and thousands of such examples. Majority of these metabolic reactions do not occur in isolation but are always linked to some other reactions. In other words, metabolites are converted into each ‘other in a series of linked reactions called metabolic pathways. These metabolic pathways are similar to the automobile traffic in a city. These pathways are either linear or circular. These pathways criss- cross each other, i.e., there are traffic junctions. Flow of metabolites through metabolic pathway has a definite rate and direction like automobile traffic. This metabolite flow is called the dynamic state of body constituents. What is most important is that this interlinked metabolic traffic is very smooth and without a single reported mishap for healthy conditions. Another feature of these metabolic reactions Is that every chemical reaction is, a catalysed reaction. There is no uncatalysed metabolic conversion in living systems. Even CO, dissolving In water, a physical process, is a catalysed reaction in living systems. The catalysts which hasten the rate of a given metabolic conversion are also proteins. These proteins with catalytic power are named enzymes. 9.4.1 Metabolic basis for living organisms. Metabolic pathways can lead to - a more complex structure from a simpler structure (for example. acetic acid becomes cholesterol or sucrose formation from water and CO, in mesopyll) or lead to a simpler structure from a complex structure (for example, glucose becomes lactie acid in our skeletal muscle). The former cases are called biosynthetic pathways or anabolic path ways. The latter constitute degradation and hence are called catabolic pathways. Anabolic pathways, as expected consume energy. Assembly of a protein reactions results from amino acids requires energy input. On the ‘other hand, catabolic pathways lead to the release of energy. For example, when glucose is degraded to lactic acid in our skeletal muscle, energy is liberated. This metabolic pathway from glucose t0 lactic acid which occurs in 11 metabolic steps 1s called glycolysis. Living organisms have learnt to trap this energy liberated during degradation and store it in the form of chemical bonds. As and when needed, this bond energy is utilised for biosynthetic, osmotic and mechanical work that we perform. The most important form of energy currency in living systems is the bond energy in a chemical called adenosine triphosphate (ATP). 9.4.2 The living state At this level, you must understand that the tens and thousands of chemical compounds in a living organism, otherwise called metabolites, or bio- molecules, are present at concentrations characteri- stic of each of them. For example, the blood concentration of glucose in a normal healthy individual is 4.5-5.0 mM, while that of hormones would be nanograms/ mL. The most important fact of biological systems is that all living organisms exist in a steady-state characterised by concentrations of each of these biomolecules. These biomolecules are in a metabolic flux. Any ‘chemical or physical process moves spontaneously to equilibrium. The steady state is an equilibrium state One should remember from physics that systems at equililbrium cannot perform work. As living organisms work continuously, they cannot afford to reach equilibrium. Hence the living state is 2 non-equilibrium steady-state to be able to perform work; living process is a constant effort to prevent falling into equilibrium state. This is achieved by energy input. Metabolism provides a mechanism for the production of energy. Hence the living state and metabolism are synonymous. Without metabolism there cannot be a living state. SINTRODUCTION ‘Almost all enzymes are protcins. There , some nucleic acids that behavelike enzy These are called ribozymes. Enzymes are defin.., as biocatalysts synthesized by living ce) Dixon and Web defined enzyme as “a protein wy, catalytic properties due to its power of speci * activation ‘One can depict an enzyme by a line dia; ‘An enzyme like any protein has a primary structur. i.e., amino acid sequence of the protein. An enzyn like any protein has the secondary and the tertiar, structure. When you look at a tertiary structure (Fig. 9.5.1). | ee Fi You will notice that the backbone of the protein chain folds upon itself, the chain criss-crosses itsclt and hence, many crevices or pockets are made ‘One such pocket is the ‘active site’. An active site of an enzyme is a crevice or pocket into which the substrate fits. Thus enzymes, through their active site, catalyse reactions at a high rate, Enzyme catalysts differ from inorganic catalysts in man) ways, but one major difference needs mention Inorganic catalysts work efficiently at high temperatures and high pressures, while enzymes get damaged at high temperatures (say above 40°C). However, enzymes isolated from organisms who normally live under extremely high temperatures (Eg. hot vents and sulphur springs are stable and retain their catalytic power even 3! high temperatures (upto 80°-90°C). Thermal stability is thus an important quality of such enzymes isolated from thermophilic organisms |. 9.5.1 Structure of Protein gps cHEMICAL REACTion, J compounds undergo phy.ic, Ca sctions. In a physical proce, ee Aa bre shape or in states of m, merging of bonds. In a chemical req pen or new bonds are formed BAOH): +350, BAS0,+2H.0 i, ganic chemical reaction. Hydrolysis of star a rch is an organic chemical rege - io glveose Bi = cal reaction, Rate agasenction is expressed as 2 Rate of catalysed reactions is hj pared to Fate of uncatalysed reactig af prysieal and chemical processes are influences py wemperature among other factors. The rate ‘or decreases to half for every 10°C ¢ hange in temperature. In catalysed reactions enzymes secelerate the rate by about 10 million times €0, + H,0 > H,Co, In the absence of any enzyme this reaction is vey slow, with about 200 molecules of H,Co, teing formed in an hour. However, by using the enzyme present within the cytoplasm called catbonie anhydrase, the reaction speeds dramatically with about 600,000 molecules being formed every second. The enzyme has accelerated the reaction rate by about 10 million times. A multi step chemical reaction when every step is catalysed by same or different enzymes is called as metabolic pathway. A particular metabolic pathway with one or two additional reactions gives fise (6 different types of metabolic end products Glucose forms pyruvic acid under aerobic condi- tions, lactic acid under anaerobic conditions of musele cells, and ethanol under anaerobic in yeast cells. Hence in different different products are possible and 88 there may alter without iction, bonds igh when ns. Rates. is an organic catalyst. It remains jing a reaction, catalysed by it. It up rate of reaction and never ilibrium of a reaction J | Specificity Enzymes are reac | catalyses only one Particular substr Sucrase, catalyses and no other subst Reversibility Most of th: | Th tion specific. One enzyme Specific reaction or acts on a ‘ate, For example, enzyme the hydrolysis of only sucrose trate. '¢ enzymes are reversible in their action *¥ an speed up a particular reaction either in | forward or in backward direction, For example, | | {Re enzyme aldolase converts a hexose into | (Wo tioses, and the same enzymes forms one hexose from two trioses. Active in minute quantity Enzymes are very active in extremely small | uantities. The number of substrate molecule | Converted into products by one molecule of an | enzyme in one minute is called turn over number (TON). Typical tum over number varies form 10? to 10° sec~!. Enzyme efficiency is very | low in Lysozyme Enzyme TON Sucrase 10,000 Urease 2000,00, Catalase 5 million Carbonic anhydrase 36 million ‘The tum over number of the enzyme is dependent upon the number of active sites present on one enyzme molecule. Therefore catalytic efficiency of fan enzyme is dependent on number of active sites. Molecular Weight Enzymatic proteins are substances of high molecular weight. Peroxidase, has molecular weight of 40,000 Daltons and Catalase- 2,50,000 Daltons pepsin 35,000 daltons, urease 483,000 daltons, Colloidal nature They are collodial in nature due to which they occupy a large surface area during the process of catalysis. Sensitivity Enzymes are heat sensitive, They also show sensitivity to pH. 9.8 HOW DO ENZYMES BRING ABOUT SUCH HIGH RATES OF CHEMICAL CONVERSIONS? yes a little To understand this we should study enzym more. We have already understood the idea of an ‘active site’. The chemical or metabolic conversion refers to a reaction. The chemical which is converted ssubstrate’. Hence into a product is called a dimensional enzymes, i.e, proteins with three structures including an ‘active sit substrate (S) into a product (P). Symbol can be depicted as: SsoP It is now understood that the substrate ‘S’ has to bind the enzyme at its ‘active site’ within a given cleft or pocket. The substrate has to diffuse towards the ‘active site’. There is thus, an obligatory formation of an “ES*complex. E stands for enzyme. This complex formation is a transient pheno- menon. During the state where substrate is bound to the enzyme active site, a new structure of the substrate called transition state structure is formed. Very soon, after the expected bond breaking/ making is completed, the product is released from the active site. In other words, the structure of substrate gets transformed into the structure of product(s). The pathway of this transformation must g0 through the so-called transition state structure. ‘There could be many more ‘altered structural states’ between the stable substrate and the product. Implicit in this statement is the fact that all other intermediate structural states are unstable. Stability is something related to energy status of the molecule or the structure. Hence, when we look at this pictorially through a graph it looks like something as in (Fig. 9.8.1). The y-axis represents the potential energy content. The x-axis represents the progression of the structural transformation or states through the “transition state’. You would notice two things. The energy level difference between S and P. If “P’ is at a lower level than ‘S’, the reaction is an exothermic reaction. One need not supply energy (by heating) in order to form the product. _ convert a ly, this Fig. 9.8.1 A change in the free energ content of the enzyme catalysed react Enzyme lowers the energy barrier However, whether it is an exothermic or sp neous reaction or an endothermic or ener *$” has to go throug! requiring reaction, the higher energy state or transition state difference in average energy content of *S" fr that of this transition state is called ‘activa energy’. Enzymes eventually bring down ¢h energy barrier making the transition of °S’ ( more easy. 9.8.1 Nature of Enzyme Action Each enzyme (E) has a substrate (S) binding site its molecule so that a highly reactive enzym substrate complex (ES) is produced. This comp! is short-lived and dissociates into its product(s and the unchanged enzyme with an intermedi formation of the enzyme-product complex (EP The formation of the ES complex is essen: for catalysis. E+S—3ES —> [EP] —> E+P Lock and Key theory or Template mode! This theory was proposed by Emil Fisher ' explain specify of enzymes. ‘He explain that interacation between en2)"° and substrate is comparable to interact between lock and key. According to‘ concept, the catalytic site of the enzyme it sel! * comple-mentary in shape to that of subst''* (Fig. 9.8.2). 66€-€ Eva Fig. 9.8.2 Lock and Key theory Induced fit theory This theory was proposed by Koshland (1958) put-not in the free-enzyme. In induced fit hypothesis active Sile’ is flexible allowing the enzyme to bind with substrate which is thereby, bringing catalytic sites and re. groups together. A conformational change takes place in the enzyme during the binding of the substrate, which results in the required matching of structures. (Fig. 9.8.3) ‘Substrate rigid, acting Enzyme-substrate complex Active site > Enzyme Fig. 9.8.3 Induced-fit model by Koshiand ‘The active site of the enzyme is flexible and ‘adjusts its conformation to fit the substrate molecule The catalytic cycle of an enzyme action can be described in the following steps: 1. First, the substrate binds to the active site of the enzyme, fitting into the active site 2. The binding of the substrate induces the enzyme to alter its shape, fitting more tightly around the substrate. 3. The active site of the enzyme, now in close Proximity of the substrate breaks the chemical bonds of the substrate and the new enzyme- Product complex is formed. The enzyme releases the products of the reaction and the free enzyme is ready to bind 0 another molecule of the substrate and run through the catalytic cycle once again. 9.9 FACTORS AFFECTING ENZYME ACTIVITY y of an enzyme can be affected by a change in the conditions which can alter the tertiary structure of the protein. These include temperature, PH, change in substrate concentration or binding of specific chemicals that regulate its activity. Effect of pH The enzymes function within a moderate pH range. Changes in pH affect enzymatic activity by altering the ionization of the enzyme or substrate This in turn influences enzyme conformation and substrate binding. Lowering the pH (increasing the Proton concentration) causes protons to bind to various enzyme side chains and substrate groups. Extreme changes in pH usually lead to enzyme inactivation. The activ Optimum enzyme activity is typically observed around pH equal to seven which is close to the PH within cells. Certain digestive enzymes prefer 4 distinctly acidic or alkaline environment (trypsin between 7.8 and 8.7). Enzymes like pepsin are active at pH values 1.5 and 2.5. The enzyme activity declines gradually on either side of the optimum. Excessive acidity or alkalinity renders enzymes inactive (Fig. 9.9.1 & Table 9.9.1) 3 2 pH Fig. 9.9.1 Effect of pH on enzyme activity When pH is plotted against enzyme activity, with the increase in pH value enzymatic activity also increases upto a certain range and gives a bell shaped curve. The peak of the curve gives optimum pH. | 9) Lipase (pancreas) ‘Temperature can also influence enzyme activity by altering conformation. Most enzymes are thermo. labile. Enzymes are most active within the optimum range of liting body temperature 25 to 48°C At a temperature below freezing point, an enzyme is inactivated Food may be preserved for long in a frozen State because neither microbia enzymes in the food can act at a 10 cause its spoilage. Ata high temperature (above 60°C) enzymes may be destroyed or denatured. Papain is used as meat tenderizer because it Continues its action even at the high temperature of cooking I enzymes or lower temperature Enzymes from certain blue -green algae that live in hot spring can function efficiently at {emperature exceeding 90°C. But enzymes from arctic micro organisms are active at temperature Rear freezing (Fig. 9.9.2) Temperature Sie. 9.8.2 Efiect of T on the enzyme activity (BOTAN) The ratio of rate of a reaction at temperatur: 10°C to that of °c is known as temperai, coefficient (Q,,). If its value is “2” that means , of reaction is doubled when temperature increased by 10°C. Concentration of Substrate With the increase in substrate concentration Velocity of the enzymatic reaction rises at first reaction ultimately reaches a maximum velo, (Vmax) which is not exceeded by any further , in concentration of the substrate. This is beca, the enzyme molecules are fewer than the subs, molecules and after saturation of these moi there are no free enzyme molecules to bind » the additional substrate molecules (Fig. 9. Temperature coefficient lecul Ko (Ss) Fig. 9.9.3 Enzyme catalysed reaction Showing hyperbolic curve Michaelis-Menten, constant The Michaelis - Menten equation may b¢ VouslS} v,=NaulS) Sneed ame where V, = observed initial velocity at given substrate concentration (S}, K_ = Michaciis constant (Moles per liter) and V__. = maximuin velocity at saturating concentration of substrate K,. value can determined by Varying the Substrate concentrations and than identifying Substrate concentration at which initial velocity is half maximal i.e a Significance of K,, (0) Reaction It indicates the affinity between enzyme ang| Subsale - substrate. io. K,, value ranges between 10- — 19-6 Molt. Enzyme = pe @ Aan enzyme having high catalytic efficiency wit Enzyme binds Enzyme releases have low K,, value and high tun over number (hin substrate Products : rd E ao Actnve K,, Tepresents an inverse measure of 2 enzyme's affinity to the substrate. The smaller | © the value of Km, the more strongly the enz Enzyme binds —_ Inhibitor competes yme wnister botete binds to the substrate. The enzyme with a high a K., has lower affinity for its substrate, Fig. 9.10.1 Competetive inhibition 2910 ENZYME INHIBITION Se ee ee eee) The activity of an enzyme is also sensitive to the i ides presence of specific chemicals that bind o the | iaeptroate 8 alee [sons enzyme. When the binding of the chemical shuts off enzyme activity, the process is called inhibition and the chemical is called an inhibitor. Foreign eae substances (other than substrate molecules) can block the catalytic effects of enzymes, These substances are called enzyme inhibitors. The dihydrofolic acid manner in which they bring about such action is called inhibition. There are three different types of dihydrofolate 3 — inhibitions. reduclase Competitive inhibition ‘These inhibitiors resemble the structure of substrate tetrahydrofolic acid and are also called as “substrate analogues”. 1 Competitive inhibitors bind to active site of enzyme there by decreasing the interaction folic acid between substrate and enzyme. Rate of competitive » 1 inhibition can be decreased by increasing the Pementi on of substrate. Sulfa drugs competitively inhibit above enzyme and act as anti micorbial agent Eg. Malonic acid is a competitive inhibitor for succinic dehydrogenase enzyme of TCA cycle. Non competitive inhibition As both succinic and malonic acid are similar in Inhibitors which do not resemble the substrate in Structure to great extent succinic dehydrognase structure but decrease the activity of the enzyme enzyme can-not differentiate between substrate and are called non competitive inhibitors, and this competitive inhibitor. These type of inhibitors are process is called non competitive inhibition. They Useful in treating cancers as well as in controlling bind to the enzyme at any place other than active bacterial pathogens (Fig. 9.10.1). site and renders it inactive. The shape of the active site is changed. Hence the substrate molecule cannot occupy the active site (Fig, 9.10.2) Eg, Salts of heavy metals like copper, mercury silver and cyanides (toxins). Other than active site biocked by Inhibitor Fig, 9.10.2 Non competitive inhibition Inhibitor All ic modulators Accumulation of end products after a series of reactions will decrease the rate of the process, Let us consider a compound A is converted to D via B and C. When ‘D’ gets accumulated, it combines with first enzyme to inhibit the formation of B. This is known as feed back inhibition. For example, when the accumulation of Glucose 6 phosphate occurs, it inhibits the enzyme hexokinase as described in (Fig. 9.10.3), ACTIVE ENZYME INACTIVE ENZYME Alosteeric transition % Inhibitor ‘Substrate Substrate x A+B} C+D GLUCOSE Be DeeoeNace PINHIBITS ADP GLUCOSE 6-PHOSPHATE Fig. 9.10.3 Feedback inhibition by the accumulation of end product which changes the substrate site on the hexokinase ‘The term allosteric (allos = other, steros = site) has been introduced by Nobel laureates Jacot, & Monad to denote an enzyme site other than active site. Enzymes which posses allosteric site, are known as allosteric enzymes. The binding Of substance to allosteric site may stimulate or inhibit enzyme action. The substances that reduce the activity of an enzyme by binding at allosteric site are known as allosteric inhibitors In feedback inhibition, the end product acts inhibitor of the enzyme of first reaction and is a Part of homeostatic control of metabolism. | Effect of enzyme inhibition Enzyme Inhibition S| 3 Competitive 8 inhibitor 3 = 2 ~~ Noncompetitive & inhibitor ‘Substrate concentration Fig. 9.10.4 9.11 CLASSIFICATION AND NOMENCLATURE OF ENZYMES Thousands of enzymes have been discovered isolated and studied. Most of these enzymes have been classified into different groups based on the type of reactions they catalyse. Enzymes are divided into 6 classes each with 4-13 subclasses and named accordingly by a four-digit number, Oxidoreductases/dehydrogenases : Enzymes which catalyse oxidoreduction between two substrates S and S' e.g, S reduced + S' oxidised — § oxidised + S' reduced Bs, Mane + NAD* pois Oxaloacetate + NADH + H” ransferases: Enzymes cata) of a group, G (other than hydrogen of substrate S and S' e.g, $-G+S —> sis Eg. Glucose+ ATP Sing a transfer ) between a pair Hexo kinase Glucose 6 phosphate + ADP Hydrolases: Enzymes cata} Ysing hydrolysis of ester, ether, peptide, glycosigi le, C-C, C-halide or PN bonds. Eg. Fructose 1.6 bisphosphate + H,0 Fructose! isphosphatase fructose 6 phosphate + Morganic phosphate Lyases: Enzymes that catal: groups from substrates by mecha; hydrolysis leaving ‘double bonds x Y ie C—C—»x-yicic Eg. Arginosuccinicacid—Awinowcsine Arginine+Fumaricacid Tsomerases: Includes all enzymes catalysing interconversion of optical, geometric or positional isomers. - Eg. Ribose Sphosphate—S

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