PAPER No. : 16 Bioorganic and biophysical chemistry MODULE No. : 25 Coenzyme-I Coenzyme A, TPP, B12 and biotin

Transcription

1 Subject Paper No and Title Module No and Title Module Tag 16, Bio organic and Bio physical chemistry 25, Coenzyme-I : Coenzyme A, TPP, B12 and CHE_P16_M25

2 TABLE OF CONTENTS 1. Learning Outcomes 2. Introduction 3.Biotin 3.1 Structure 3.2 Function 3.3 Mechanism of carboxylation 4. Coenzyme A 4.1 Structure 5. B12 Coenzyme 5.1 Structure 6. Thiamine pyrophosphate 6.1 Introduction and structure 6.2 Role of thiamine phosphate as a Coenzyme 6.3 Thiamine pyrophosphate in the pyruvate dehydrogenase reaction. 7 Summary

3 1. Learning Outcomes After studying this module, you shall be able to Know about the structure and functions of Biotin, Coenzyme A, TPP (Thiamine Pyrophosphate) and B 12 Learn mechanism of function of these co-enzymes Analyze role of thiamine phosphate as a coenzyme 2. Introduction Coenzymes are non-protein molecules that function as essential parts of enzymes. Coenzymes often serve as special reagents needed for reactions that would be difficult or impossible using only simple acid-base catalysis. In many instances, they also serve as carriers alternating catalysts that accept and donate chemical groups, hydrogen atoms, or electrons. They are often complex organic structures that cannot be synthesized by some organisms - mammals in particular. The water soluble vitamins, those usually referred to as the vitamin B-complex, are metabolic precursors of a number of coenzymes. That is why vitamins are so important in metabolism. Coenzymes are considered under different groups depending on types of reaction they carry out. In this we will consider those group of compounds, which are derivatives of vitamins that while in the active site of enzyme, alter the structure of a substrate in a way that permits it to react more readily, or we can call them group transferring coenzymes. Biotin, Coenzyme A, TPP (Thiamine Pyrophosphate) and B Biotin 3.1 Structure Biotin is an imidazole ring fused with thiophene ring with valeryl side chain.

4 Fig1. Structure of Biotin is widely distributed in food as biocytin, which is bound to amino group of lysine. Fig2. Structure of biocytin 3.2 Function This water soluble B group vitamin participates in transfer of carboxylic group. Two important enzymes carrying out carboxylation reaction are i. Pyruvate carboxylase ii. Acetyl CoA carboxylase

5 3.3 Mechanism of carboxylation A holo carboxylase synthetase catalyses the attachment of to ε amino group of lysine residue of carboxylase enzyme to form biocytin, in an ATP dependent reaction. Fig3. Mechanism of carboxylation Biocytin as a part of carboxylase enzyme then catalyzes the transfer of CO 2 to the substrate. It first attaches to activated CO 2 [from HCO - 3] to form carboxylated. This then transfers carboxylate to the substrate. Enzyme bound acts as a carrier for CO 2 in pyruvate carboxylase reaction. The dependent enzymes require ATP which is converted to ADP during reaction. Pyruvate carboxylase is a tetrameric protein carrying four moleculeus of, each bound covalently through an amide bond involving ε amino group of lysine residue. Pyruvate carboxylase operates by a two-step mechanism.

6 Fig4. Enzyme bound acts as a carrier for CO2 in pyruvate carboxylase reaction Starting with an ATP dependent carboxylation of cofactor to give n-carboxy. This activated derivative then transfers the carboxyl directly to pyruvate. 4. Coenzyme A Coenzyme A is an acyl activating enzyme derived from vitamin pantothenic acid. Panthotheric acid is pentoic acid and β alanine joined together in a amide linkage

7 Coenzyme A can be divided into two components, adenosine 3, 5 diphosphate and pantotheine, which is formed the combination of pantothenic acid and βmercaptoethylamine. 4.2 Structure Fig5. Structure of Coenzyme A The SH group of thioethanol amine moeity is an active group acting as carrier and coenzyme A is abbreviated as CoA or CoASH. The rest of molecule provides enzyme binding site. In acylated derivatives, such as acetyl-coenzyme A, the acyl group is linked to thiol group of form an energy rich thio ester.

8 CoA reacts with many compounds to form important derivatives such as: Acetyl CoA Succinyl CoA HMG CoA Fatty acyl CoA Acetyl CoA and succinyl CoA are important intermediates at cross roads of many metabolic pathways. Acyl coa is formed as an intermediate both in fatty acid biosynthesis and oxidation. HMG CoA is an important intermediate involved in ketogenesis as well as cholestrol biosynthesis we designate the acylated forms of coenzyme A as or acyl coa, and unacylated as CoA-SH. While CoA was discovered as the "acetylation coenzyme. It has far more general function. It is required in the form of acetyl CoA, to catalyze the synthesis of citrate in citric acid cycle. It is essential to the β oxidation of fatty acids and carries propionyl and other acyl groups in a great variety of other metabolic reactions. The acyl group [such as acetyl or aceto acetyl group] is attached to the CoA through a thio ester linkage to the β- mercaptoethylamine moiety. Acyl groups are covalently linked to thiol group, forming thioesters. Because of their relatively high standard free energies of hydrolysis, thioesters have high acyl group transfer potential and cars donate their acyl group of variety of acceptor molecules. The acyl group attached to coenzyme A may thus be thought of as "activated" for group transfer. 5. Coenzyme B12 Coenzyme B 12 is the cofactor form of vitamin B 12. This vitamin is unique among all the vitamins in that it not only contains complex organic molecule but an essential trace element Cobalt. The structure of vitamin B 12 is shown in.

9 Fig6. Structure of Coenzyme B12 The tetra pyrrole ring system, called a corrin ring similar to porphyrin ring of heme compounds. The cobalt is also linked to heterocylic base 5, 6 dimethy l benzimidazole (DMB) ribonucleotide (fig.) In the vitamin as it is isolated sixth coordination position of cobalt is occupied by cyanide ion, but this ion is introduced during isolation.

10 The vitamin as it exists in tissues contains either water or a hydroxy moiety at this site because of presence of cobalt and many amide nitrogen's B 12 compounds are called cobamides or cobalamins. B 12 derivatives are also named in terms of group occupying the sixth coordination position. Thus, the vitamins as isolated is cyanocobalamin, and intracellular forms are aquocobalamin or hydroxocobalami. 5.1 Co-enzyme form of B12 Two active forms of vitamin B 12 are involved in metabolism. These are (a) methyl cobalamin and b) 5 - deoxy adenosyl cobalamine also called cobamide coenzyme only two reactions use vitamin B 12 as coenzymes. Conversion of homo cysteine to Methionine, Methyl cobalamine is the coenzyme in this reaction. Conversion of methyl malonyl CoA to succinyl CoA and 5 - deoxy adenosyl cobalamine is used as coenzyme. Function of 5 - deoxyadenosylcobalamin involves isomerizations involving exchange of carbon bound hydrogen with another carbon bound functional group as shown here methyl malonyl CoA mutase catalyzes reaction of this type

11 6. Thiamine pyrophosphate 6.1 Introduction and structure Thiamine was the first of B vitamins to be identified. It is also called as vitamin B 1. The vitamin is structurally complex, made up of 2, 5 dimethyl 6-amino pyrimidine joined to 4 methyl, 5 hydroxythyl thiozole by a methylene linkage. But its conversion to the coenzyme form, thiamine pyrophosphate, or TPP involves simply and ATP dependent pyrophosphorylation. [Chemical nature] Thiazole ring of TPP is the functional part of coenzyme. Dietary thiamine after absorption is converted to thiamine pyrophosphate by thiamine kinase in most tissues in our body using ATP.

12 6.2 Role of thiamine phosphate as a Coenzyme Thiamine pyrophosphate participate as a coenzyme in following reactions: Oxidative decarboxylation of pyruvate, α ketoglutarate and branced chain amino acids Α keto acid dehydrogenase reaction of branched amino acid Transketolase reaction of HMP shunt pathway Thiamine pyrophosphate is the coenzyme for all decarboxylations of α-keto acids. The mechanism shown in Figure 14.6 for pyruvate decarboxylation is involved in all of these reactions. Note that TPP contains two heterocyclic rings a substituted pyrimidine and a thiazole. Recent NMR studies have shown that both rings participate in the formation of a reactive carbanion at C-2 of the thiazole ring-the carbon atom between the nitrogen and the sulfur. As shown in the following diagram, a glutamate carboxyl group in the enzyme attracts a proton. Linked to N-1 of the pyrimidine, which in turn increases the basicity of the amino group, facilitating the deprotonation of C-2 of the thiazole ring. This carbon forms a carbanion (step 1 in Figure) which in turn can attack the carbonyl carbon of a-keto acids, such as pyruvate, giving an addition compound (step 2). The addition compound undergoes nonoxidative decarboxylation (step 3) with the thiazole ring acting as an elect stabilized eneamine. Protonation (step 4) give hyde or more accurately, hydroxyethyl-tpp.

13 6.3 Thiamine pyrophosphate in the pyruvate dehydrogenase reaction. Thiamine pyrohphosphate (TPP) is the coenzyme for the pyruvate dehydrogenase reaction and other nonoxidative de-carboxylations of α-keto acids. The key reaction (step 2) is attack by the carbanion of TPP on the carbonyl carbon of pyruvate and is followed by nonoxidative decarboxylation of the coenzyme - bound pyruvate (step 3) the electron pair remains with the ring nitrogen. In step 4, the two-carbon fragment (red) bound to TPP extracpts a proton from pyruvate decarboxylase (not shown), generating a hydroxythyl group. This fragments remains at the aldehyde oxidation level. 7. Summary Four enzymes which are involved in transfer of groups are vit B 12,, TPP, and coenzyme A. Biotin participates in the transfer of carboxylic groups. Thiamine pyrophosphate or thiamine TDP is the coenzyme responsible for the transfer of aldehyde and glyoxal group, and it is derived from vitamin B, by phosphorylation.

14 Coenzyme A is derived from vitamin pantothenic acid. This is abbreviated as CoA. It is also called as acetylation coenyzme Coenzyme B 12 is the cofactor form of vit B 12 and involved in isomerisation reaction and methyl group transfer.