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 2

1.4 The commercial importance of enzymes in biosynthesis and biotechnology 3

1.5 The importance of enzymes as targets for drug discovery 5

2 All Enzymes Are Proteins 7

2.1 Introduction 7

2.2 The structures of the L--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 19

2.9 Membrane-associated Enzymes 20

2.10 Glycoproteins 21

3 Enzymes are Wonderful Catalysts 25

3.1 Introduction 25

3.2 A thermodynamic model of catalysis 27

3.3 Proximity effects 28

3.4 The importance of transition state stabilisation 31

3.5 Acid/base catalysis in enzymatic reactions 34

3.6 Nucleophilic catalysis in enzymatic reactions 37

3.7 The use of strain energy in enzyme catalysis 41

3.8 Desolvation of substrate and active site nucleophiles 42

3.9 Catalytic perfection 44

3.10 The involvement of protein dynamics in enzyme catalysis 44

4 Methods for Studying Enzymatic Reactions 47

4.1 Introduction 47

4.2 Enzyme purification 47

4.3 Enzyme kinetics 49

4.4 The stereochemical course of an enzymatic reaction 55

4.5 The existence of intermediates in enzymatic reactions 61

4.6 Analysis of transition states in enzymatic reactions 64

4.7 Determination of active site catalytic groups 67

5 Hydrolytic and Group Transfer Enzymes 72

5.1 Introduction 72

5.2 The peptidases 73

5.3 Esterases and lipases 85

5.4 Acyl transfer reactions in biosynthesis (coenzyme A) 86

5.5 Enzymatic phosphoryl transfer reactions 88

5.6 Adenosine 5-triphosphate (ATP) 93

5.7 Enzymatic glycosyl transfer reactions 95

5.8 Methyl group transfer: use of S-adenosyl methionine and tetrahydrofolate coenzymes for one-carbon transfers 99

6 Enzymatic Redox Chemistry 108

6.1 Introduction 108

6.2 Nicotinamide adenine dinucleotide-dependent dehydrogenases 110

6.3 Flavin-dependent dehydrogenases and oxidases 115

6.4 Flavin-dependent mono-oxygenases 120

6.5 CASE STUDY: Glutathione and trypanothione reductases 122

6.6 Deazaflavins and pterins 126

6.7 Iron-sulphur clusters 127

6.8 Metal-dependent mono-oxygenases 128

6.9 -Ketoglutarate-dependent dioxygenases 131

6.10 Non-heme iron-dependent dioxygenases 133

7 Enzymatic Carbon-Carbon Bond Formation 139

7.1 Introduction 139

Carbon-carbon bond formation via carbanion equivalents 140

7.2 Aldolases 140

7.3 Claisen enzymes 144

7.4 Assembly of fatty acids and polyketides 146

7.5 Carboxylases: Use of biotin 150

7.6 Ribulose bisphosphate carboxylase/oxygenase (Rubisco) 151

7.7 Vitamin K-dependent carboxylase 153

7.8 Thiamine pyrophosphate-dependent enzymes 155

Carbon-carbon bond formation via carbocation intermediates 158

7.9 Terpene cyclases 158

Carbon-carbon formation through radical intermediates 162

7.10 Phenolic radical couplings 163

8 Enzymatic Addition/Elimination Reactions 170

8.1 Introduction 170

8.2 Hydratases and dehydratases 171

8.3 Ammonia lyases 175

8.4 Elimination of phosphate and pyrophosphate 177

8.5 CASE STUDY: 5-Enolpyruvyl shikimate 3-phosphate (EPSP) synthase 180

9 Enzymatic Transformations of Amino Acids 185

9.1 Introduction 185

9.2 Pyridoxal 5-phosphate-dependent reactions at the -position 185

9.3 CASE STUDY: Aspartate aminotransferase 189

9.4 Reactions at the - and -positions of amino acids 192

9.5 Serine hydroxymethyltransferase 195

9.6 N-Pyruvoyl-dependent amino acid decarboxylases 195

9.7 Imines and enamines in alkaloid biosynthesis 196

10 Isomerases 200

10.1 Introduction 200

10.2 Cofactor-independent racemases