Introduction to Enzyme and Coenzyme Chemistry

Enzymes are giant macromolecules which catalyse biochemical reactions. They are remarkable in many ways. Their three–dimensional structures are highly complex, yet they are formed by spontaneous folding of a linear polypeptide chain. Their catalytic properties are far more impressive than synthetic catalysts which operate under more extreme conditions. Each enzyme catalyses a single chemical reaction on a particular chemical substrate with very high enantioselectivity and enantiospecificity at rates which approach “catalytic perfection”. Living cells are capable of carrying out a huge repertoire of enzyme–catalysed chemical reactions, some of which have little or no precedent in organic chemistry. The popular textbook Introduction to Enzyme and Coenzyme Chemistry has been thoroughly updated to include information on the most recent advances in our understanding of enzyme action, with additional recent examples from the literature used to illustrate key points. A major new feature is the inclusion of two–colour figures, and the addition of over 40 new figures of the active sites of enzymes discussed in the text, in order to illustrate the interplay between enzyme structure and function. This new edition provides a concise but comprehensive account from the perspective of organic chemistry, what enzymes are, how they work, and how they catalyse many of the major classes of enzymatic reactions, and will continue to prove invaluable to both undergraduate and postgraduate students of organic, bio–organic and medicinal chemistry, chemical biology, biochemistry and biotechnology.

Preface ix Representation of Protein Three–Dimensional Structures x 1 From Jack Beans to Designer Genes 1 1.1 Introduction 1 1.2 The discovery of enzymes 1 1.3 The discovery of coenzymes 3 1.4 The commercial importance of enzymes in biosynthesis and biotechnology 3 1.5 The importance of enzymes as targets for drug discovery 6 2 All Enzymes Are Proteins 7 2.1 Introduction 7 2.2 The structures of the L–α–amino acids 7 2.3 The primary structure of polypeptides 9 2.4 Alignment of amino acid sequences 11 2.5 Secondary structures found in proteins 12 2.6 The folded tertiary structure of proteins 15 2.7 Enzyme structure and function 17 2.8 Metallo–enzymes 20 2.9 Membrane–associated enzymes 21 2.10 Glycoproteins 23 3 Enzymes Are Wonderful Catalysts 26 3.1 Introduction 26 3.2 A thermodynamic model of catalysis 28 3.3 Proximity effects 30 3.4 The importance of transition state stabilisation 32 3.5 Acid/base catalysis in enzymatic reactions 36 3.6 Nucleophilic catalysis in enzymatic reactions 40 3.7 The use of strain energy in enzyme catalysis 44 3.8 Desolvation of substrate and active site nucleophiles 45 3.9 Catalytic perfection 46 3.10 The involvement of protein dynamics in enzyme catalysis 47 4 Methods for Studying Enzymatic Reactions 50 4.1 Introduction 50 4.2 Enzyme purification 50 4.3 Enzyme kinetics 52 4.4 The stereochemical course of an enzymatic reaction 59 4.5 The existence of intermediates in enzymatic reactions 64 4.6 Analysis of transition states in enzymatic reactions 68 4.7 Determination of active site catalytic groups 71 5 Hydrolytic and Group Transfer Enzymes 77 5.1 Introduction 77 5.2 The peptidases 79 CASE STUDY: HIV–1 protease 90 5.3 Esterases and lipases 92 5.4 Acyl transfer reactions in biosynthesis (coenzyme A) 93 5.5 Enzymatic phosphoryl transfer reactions 95 5.6 Adenosine 5’–triphosphate (ATP) 101 5.7 Enzymatic glycosyl transfer reactions 102 5.8 Methyl group transfer: use of S–adenosyl methionine and tetrahydrofolate coenzymes for one–carbon transfers 107 6 Enzymatic Redox Chemistry 115 6.1 Introduction 115 6.2 Nicotinamide adenine dinucleotide–dependent dehydrogenases 117 6.3 Flavin–dependent dehydrogenases and oxidases 122 6.4 Flavin–dependent mono–oxygenases 128 6.5 CASE STUDY: Glutathione and trypanothione reductases 129 6.6 Deazaflavins and pterins 133 6.7 Iron–sulphur clusters 135 6.8 Metal–dependent mono–oxygenases 136 6.9 α–Ketoglutarate–dependent dioxygenases 140 6.10 Non–heme iron–dependent dioxygenases 141 7 Enzymatic Carbon–Carbon Bond Formation 148 7.1 Introduction 148 Carbon–carbon bond formation via carbanion equivalents 149 7.2 Aldolases 149 CASE STUDY: Fructose 1,6–bisphosphate aldolase 150 7.3 Claisen enzymes 153 7.4 Assembly of fatty acids and polyketides 156 7.5 Carboxylases: Use of biotin 158 7.6 Ribulose bisphosphate carboxylase/oxygenase (Rubisco) 161 7.7 Vitamin K–dependent carboxylase 163 7.8 Thiamine pyrophosphate–dependent enzymes 165 Carbon–carbon bond formation via carbocation intermediates 168 7.9 Terpene cyclases 168 Carbon–carbon formation through radical intermediates 173 7.10 Phenolic radical couplings 173 8 Enzymatic Addition/Elimination Reactions 181 8.1 Introduction 181 8.2 Hydratases and dehydratases 182 8.3 Ammonia lyases 187 8.4 Elimination of phosphate and pyrophosphate 190 8.5 CASE STUDY: 5–Enolpyruvyl shikimate 3–phosphate (EPSP) synthase 191 9 Enzymatic Transformations of Amino Acids 197 9.1 Introduction 197 9.2 Pyridoxal 5’–phosphate–dependent reactions at the α–position 197 9.3 CASE STUDY: Aspartate aminotransferase 201 9.4 Reactions at the β– and γ–positions of amino acids 204 9.5 Serine hydroxymethyltransferase 206 9.6 N–Pyruvoyl–dependent amino acid decarboxylases 208 9.7 Imines and enamines in alkaloid biosynthesis 208 10 Isomerases 213 10.1 Introduction 213 10.2 Cofactor–independent racemases and epimerases 213 10.3 Keto–enol tautomerases 216 10.4 Allylic isomerases 217 10.5 CASE STUDY: Chorismate mutase 219 11 Radicals in Enzyme Catalysis 225 11.1 Introduction 225 11.2 Vitamin B12–dependent rearrangements 225 11.3 The involvement of protein radicals in enzyme catalysis 229 11.4 S–adenosyl–methionine–dependent radical reactions 232 11.5 Biotin synthase and sulphur insertion reactions 233 11.6 Radical chemistry in DNA repair enzymes 234 11.7 Oxidised amino acid cofactors and quinoproteins 238 12 Non–Enzymatic Biological Catalysis 242 12.1 Introduction 242 12.2 Catalytic RNA 242 12.3 Catalytic antibodies 246 12.4 Synthetic enzyme models 251 Appendix 1: Cahn–Ingold–Prelog Rule for Stereochemical Nomenclature 258 Appendix 2: Amino Acid Abbreviations 260 Appendix 3: A Simple Demonstration of Enzyme Catalysis 261 Appendix 4: Answers to Problems 263 Index 271

“Summing Up: Recommended.  Lower–and upper–division undergraduates.”  ( Choice , 1 April 2013)