AN INTRODUCTIONTO AGRICULTURALBIOCHEMISTRY

J.M. ChesworthHonorary Lecturer in Agricultural Biochemistry and NutritionDepartment of AgricultureUniversity of Aberdeen, UK

and

Independent ConsultantMar Technical ServicesHuntly, UK

T. StuchburyLecturer in Agricultural Biochemistry and Plant PhysiologyDepartment of AgricultureUniversity of Aberdeen, UK

and

J.R. ScaifeLecturer in Agricultural Biochemistry and NutritionDepartment of AgricultureUniversity of Aberdeen, UK

CHAPMAN & HALLLondon • Weinheim • New York • Tokyo • Melbourne • Madras

CONTENTS

Preface xviii

List of abbreviations xix

PART ONE: THE CELL AND CELLULAR CONSTITUENTS 1

1 Cell structure and function 31.1 Introduction 31.2 Components of cells 3

1.2.1 Plasma membrane 31.2.2 Cytoplasm 41.2.3 The nucleoid and nucleus 61.2.4 Cell walls 61.2.5 Ribosomes 61.2.6 Endoplasmic reticulum 61.2.7 Vacuoles and specialized vesicles 71.2.8 Mitochondria 81.2.9 Chloroplasts 81.2.10 Cytoskeleton 8

1.3 Cell specialization and interaction 9

2 Water and solutions 112.1 Introduction 112.2 The ionization of water 12

2.2.1 The pH of water 122.3 What are acids and bases? 142.4 Biological systems, ionic strength and pH 14

2.4.1 Stabilization of pH by buffers 142.5 Colligative properties 15

2.5.1 Depression of freezing point 162.5.2 Osmotic pressure 162.5.3 Semipermeable membranes that allow some solutes to pass 17

3 The carbohydrates 193.1 Introduction 193.2 Structures of sugars 20

3.2.1 Optical isomers 203.3 Naming of sugars 203.4 Sugars with four carbon atoms, the tetroses 20

vi Contents

3.5 Sugars with five carbon atoms, the pentoses 213.5.1 Ring formation in sugars 223.5.2 Five- and six-membered rings 233.5.3 Ring formation is not permanent 23

3.6 Sugars with six carbon atoms, the hexoses 233.6.1 Glucose 233.6.2 Fructose 233.6.3 Other hexoses 23

3.7 Reducing and non-reducing sugars 243.8 Formation of sugar acetals 24

3.8.1 Formation of disaccharides 253.8.2 Sucrose 25

3.9 Polysaccharides 263.9.1 The storage carbohydrates - starch and glycogen 263.9.2 Structural polysaccharides in plants 293.9.3 Other polysaccharides and related compounds 32

4 Fatty acids and lipids 354.1 Introduction 354.2 Structure and occurrence of lipids 35

4.2.1 Fatty acids 354.2.2 Triacylglycerols and other acylglycerols 394.2.3 Glycerophospholipids 424.2.4 Glycosylglycerides 434.2.5 Sphingolipids 444.2.6 Terpenes and steroids 454.2.7 Waxes 47

5 Amino acids and proteins 515.1 Introduction 515.2 Amino acids 52

5.2.1 Structure of amino acids 525.3 Non-protein amino acids and related compounds 52

5.3.1 Canavanine 525.3.2 Selenium-containing amino acids 525.3.3 Mimosine 545.3.4 Lathyrogens 555.3.5 S-methyl cysteine sulphoxide (SMCO) 555.3.6 Alkaloids 55

5.4 Phenolics 575.4.1 Lignin 575.4.2 Tannins 575.4.3 Flavonoids 58

5.5 Peptide bonds 595.6 Protein function and structure 61

5.6.1 Primary protein structure 625.6.2 Secondary protein structure 625.6.3 Tertiary structure 63

Contents vii

5.6.4 Quaternary structure 645.7 Properties of proteins 66

5.7.1 Ionic strength and presence of specific ions 675.7.2 Effect of pH 685.7.3 Denaturation 685.7.4 Effect of heat 69

5.8 Prions 69

6 Enzymes 716.1 Introduction . 716.2 Types of reactions catalysed by enzymes 716.3 Mode of action of enzymes 726.4 Factors contributing to enzyme activity 73

6.4.1 Proximity of substrates at the active site 736.4.2 Environment 746.4.3 Acid-base catalysis 746.4.4 Effects on the stability of substrates and reaction intermediates 746.4.5 Formation of covalent enzyme-substrate intermediates 74

6.5 Factors affecting the rates of enzyme-catalysed reactions 766.5.1 Enzyme concentration 766.5.2 Substrate concentration 766.5.3 Temperature 796.5.4 pH 796.5.5 Presence of inhibitors 816.5.6 Presence of coenzymes 84

6.6 Allosteric enzymes 906.7 Molecular recognition 91

6.7.1 Receptors 916.7.2 Antibodies 91

7 Purines, pyrimidines and nucleic acids 957.1 Introduction 957.2 Purines and pyrimidines 957.3 Deoxyribonucleic acid (DNA) 96

7.3.1 Chemical nature of DNA 967.3.2 The DNA double helix 987.3.3 Structure of DNA in prokaryotes and eukaryotes 987.3.4 OrganelleDNA 100

7.4 Ribonucleic acid (RNA) 1007.4.1 Messenger RNA (mRNA) 1017.4.2 Transfer RNA (tRNA) 1017.4.3 Ribosomal RNA (rRNA) 102

8 Vitamins and minerals 1058.1 Vitamins in biochemistry 105

.8.1.1 Introduction 1058.1.2 Thiamin (vitamin Bj) 1068.1.3 Riboflavin (vitamin B2) 108

viii Contents

8.1.48.1.58.1.68.1.78.1.88.1.98.1.108.1.118.1.128.1.138.1.148.1.158.1.16

Nicotinic acid (niacin, formerly called vitamin B5)Pantothenic acidPyridoxine, pyridoxal and pyridoxamine (vitamin B6)BiotinFolic acidVitamin B12

Vitamin CCholineCarnitineVitamin AVitamin DVitamin EVitamin K

8.2 Minerals in biochemistry8.2.18.2.28.2.38.2.48.2.58.2.68.2.7

IntroductionCalciumPhosphorusMagnesiumSodium, chloride and potassiumSulphur and ironOther elements with known biochemical functions

9 The composition of agricultural products9.1 Introduction

9.1.1 Energy storage in animals and plants92 The composition of animals

9.2.1 Body composition9.2.2 Milk

9.3 Plant materials9.4 Principal nutrients in plants and animals

9.4.1 Proteins9.4.2 Lipids9.4.3 Carbohydrates

PART TWO: METABOLISM

10 Glycolysis10.1 Introduction10.2 Stage 1 - preparing glucose for splitting into two three-carbon units

10.2.1 Glucose phosphorylation10.2.2 Fructose and its phosphates10.2.3 Splitting of fructose-l,6-bisphosphate

10.3 Stage 2 - metabolism of the three-carbon compounds10.3.1 First oxidation step10.3.2 First energy released in the form of ATP10.3.3 Formation of pyruvate

10.4 The entry of other sugars10.4.1 Fructose

108112112114114115117118119119121124126127127127128128129129130

Contents ix

10.4.2 Galactose 14610.4.3 The entry of glycogen 146

11 The tricarboxylic acid cycle 14911.1 Introduction 14911.2 The reactions of the TCA cycle 149

11.2.1 Production of acetyl-CoA 14911.2.2 Reactions leading to the production of CO2 15011.2.3 Reactions leading back to the formation of oxaloacetate 15011.2.4 Overall reactions of the tricarboxylic acid cycle 153

11.3 Links with other metabolic pathways 45311.4 Replenishment of TCA cycle intermediates 15511.5 Conversion of propionate to glucose via the TCA cycle ' 15511.6 Regulation of the TCA cycle 15511.7 The glyoxylate cycle 156

12 Electron transport and oxidative phosphorylation 16112.1 Introduction 16112.2 The mitochondrion 16312.3 Components of the electron transport chain 163

12.3.1 Flavoproteins 16312.3.2 The iron-sulphur proteins 16412.3.3 Ubiquinone 164

12.4 The electron transport chain complexes 16412.4.1 Complex I - the NADH-dehydrogenase complex 16412.4.2 Complex II - the succinate dehydrogenase complex 16412.4.3 Complex III - the cytochrome b, ct complex 16512.4.4 Complex IV - cytochrome oxidase 165

12.5 Coupling of electron transport to ATP synthesis 16612.6 The yield of ATP 16812.7 NADH produced in the cytoplasm enters the electron transport chain via 169

shuttle reactions12.8 Regulation of oxidative phosphorylation by ADP/ATP supply 169

13 Fatty acid oxidation and lipid breakdown 17313.1 Introduction 17313.2 |3-Oxidation 174

13.2.1 Mitochondrial p-oxidation in animal tissues 17413.2.2 The reactions of p-oxidation 17613.2.3 P-Oxidation of odd-numbered acids 17613.2.4 P-Oxidation of unsaturated acids 17813.2.5 P-Oxidation in peroxisomes and glyoxisomes 17913.2.6 The formation of ketone bodies 181

13.3 a-Oxidation 18213.4 co-Oxidation 18313.5 Peroxidation of fatty acids 183

13.5.1 Chemistry of lipid peroxidation 18413.5.2 Prevention of fatty acid peroxidation 185

x Contents

13.5.3 Detection and measurement of lipid peroxidation 18513.5.4 Effects of peroxidation in living organisms 18613.5.5 Lipoxygenase and cyclo-oxygenase 188

13.6 Breakdown of lipids 18913.6.1 Triacylglycerol breakdown 19013.6.2 Phospholipid breakdown 19113.6.3 Breakdown of glycolipids 192

14 Breakdown of proteins and the oxidation of amino acids 19314.1 Introduction 19314.2 Breakdown of proteins 194

14.2.1 Digestion of proteins 19414.2.2 Protein turnover 194

14.3 Breakdown of amino acids 19414.3.1 Transamination reactions 19414.3.2 Deamination 19514.3.3 Oxidation of carbon skeletons of amino acids 19514.3.4 The fate of ammonia 19714.3.5 The urea cycle 19714.3.6 The fate of urea in ruminants 199

14.4 Precursor functions of amino acids 199

15 The pentose phosphate pathway 20115.1 Introduction 20115.2 Oxidative reactions 20115.3 Rearrangement reactions 20115.4 Importance of the pathway 20515.5 Regulation of the pathway 206

16 Fermentation pathways 20716.1 Introduction 20716.2 Anaerobic environments in agriculture 20716.3 Lactate production 207

16.3.1 Muscle metabolism 20816.3.2 Regeneration of glucose 208

16.4 Animal digestive systems 20816.4.1 Acetate formation 20916.4.2 Propionate formation 20916.4.3 Butyrate synthesis 210

16.5 Soils 21016.6 Waste treatment 21116.7 Methane production 21316.8 Dairy products 213

• 16.9 Meat 21316.10 Fermentation in herbages 21416.11 Ethanol production 214

17 Photosynthesis 21717.1 Introduction 217

Contents xi

17.2 Chloroplasts 21717.3 The light reactions 217

17.3.1 Photosynthetic pigments 21717.3.2 Light absorption 21917.3.3 Resonance energy transfer 22117.3.4 The electron transport system 22117.3.5 PhotosystemI 22217.3.6 Photosystem II 22317.3.7 Cytochrome b-f complex 22517.3.8 Plastocyanin 225

17.4 Integration of the electron transport system 22517.5 ATP production 22517.6 The dark reactions (Calvin cycle) 22617.7 Control of photosynthesis 22917.8 Photorespiration 229

17.8.1 Factors affecting rates of photorespiration 23017.9 Photosynthesis in C4 plants 23217.10 Crassulacean acid metabolism 23317.11 Herbicides and photosynthesis 234

18 Gluconeogenesis and carbohydrate synthesis 23718.1 Introduction 237

18.1.1 Starting materials 23718.1.2 Outline of the pathway 23818.1.3 Differences between gluconeogenesis and glycolysis 238

18.2 Gluconeogenesis via pyruvate 23818.2.1 The control of pyruvate production and use 238

18.3 The production of fructose-l,6-bisphosphate 23818.4 The hydrolysis of fructose-l,6-bisphosphate 240

18.4.1 The control of phosphofructokinase and fructose-l,6-bisphosphatase 24018.4.2 The fate of fructose-6-phosphate 242

18.5 The utilization of glucose-6-phosphate 24318.6 Gluconeogenesis from propionate 24318.7 The synthesis of complex carbohydrates 243

18.7.1 Disaccharides 24318.7.2 Synthesis of polysaccharides 246

19 Synthesis of fatty acids and lipids 25119.1 Introduction 25119.2 Tissue and subcellular location of fatty acid synthesis 25219.3 Source of the primary substrate - acetate 25219.4 Production of malonyl-CoA 253

19.4.1 Animals 25419.4.2 Plants 25619.4.3 Bacteria 256

19.5 Synthesis of long-chain saturated fatty acids from acetyl-CoA and malonyl-CoA 25619.5.1 Acyl carrier protein and its function 25619.5.2 The reactions of fatty acid synthesis 257

xii Contents

19.5.3 Chain length specificity of fatty acid synthetases 25919.5.4 Synthesis of branched-chain fatty acids 26019.5.5 Releaseof fatty acids from fatty acid synthetase 261

19.6 Fatty acid elongation 26119.7 Formation of unsaturated fatty acids 262

19.7.1 Desaturation of fatty acids in animals 26219.7.2 Desaturation of fatty acids in plants 26319.7.3 Essential fatty acids 263

19.8 Synthesis of triacylglycerols 26519.8.1 The 2-monoacylglycerol pathway 26519.8.2 The glycerol-3-phosphate pathway 26619.8.3 Triacylglycerol synthesis in plants 267

19.9 Phospholipids 26819.10 Glycolipid synthesis 26819.11 Synthesis of sphingolipids 27019.12 Biosynthesis of terpenes and sterols 271

19.12.1 Synthesis of mevalonic acid 27119.12.2 Conversion of mevalonic acid to squalene 273

20 Synthesis of amino acids 27920.1 Introduction 27920.2 Assimilation of nitrate 27920.3 Nitrogen fixation 281

20.3.1 Molecular biology of nitrogen fixation 28220.4 Assimilation of ammonia 28220.5 Biosynthesis of amino acids 284

20.5.1 Aromatic amino acids and related compounds 28420.5.2 Branched-chain aliphatic amino acids 285

20.6 Nutritional role of amino acids 28720.7 Herbicides and amino acid biosynthesis 289

21 The synthesis of nucleic acids and proteins 29121.1 Introduction 29121.2 Synthesis of purine and pyrimidine nucleotides 29221.3 Replication of DNA 292

21.3.1 DNA synthesis in viruses 29421.3.2 Accuracy of DNA synthesis 296

21.4 Synthesis of RNA 29621.5 The genetic code 29721.6 Protein synthesis 297

21.6.1 Amino acid activation 29721.6.2 Initiation 29921.6.3 Elongation 30021.6.4 Termination 30021.6.5 Post-translational modification of proteins 30021.6.6 Location of protein synthesis 301

21.7 Regulation of protein synthesis 30321.7.1 Regulation in prokaryotes 303

Contents xiii

21.7.2 Regulation in eukaryotes 30521.8 Protein synthesis in chloroplasts and mitochondria 30621.9 Genetic engineering 308

21.9.1 Enzymes used in DNA manipulation 30821.9.2 Isolation and synthesis of DNA 30821.9.3 Gene cloning 30921.9 A Screening techniques 31021.9.5 Restriction fragment length polymorphism (RFLP) 31021.9.6 Antisense RNA 31021.9.7 Vectors and methods for insertion of DNA into cells 31021.9.8 Site-specific mutagenesis (oligonucleotide-directed mutagenesis) 311

22 Compartments, membranes and regulation 31322.1 Cell compartments 31322.2 Lipids and membranes 31322.3 Transport across membranes 315

22.3.1 Membrane transport mechanisms 31622.3.2 Other functions of membranes 321

22.4 Principles of metabolic regulation 32222.4.1 Allosteric regulation 32322.4.2 Covalent modification 32322.4.3 Changes in the amount of enzyme 32422.4.4 Coordinated regulation of pathways 32522.4.5 Coordination of metabolism in different tissues 328

PART THREE: STRATEGIES FOR PROCESSING OF NUTRIENTS IN PLANTS 333

23 Seeds and germination 33523.1 Seeds and plant development 33523.2 Seeds as food, and agricultural commodities 33523.3 Seed composition 336

23.3.1 Seed carbohydrates 33623.3.2 Seed lipids 33723.3.3 Seed proteins 33723.3.4 Seed minerals 339

23.4 Germination 34023.4.1 Starch breakdown 34023.4.2 Beer and whisky production 34123.4.3 Protein breakdown 34223.4.4 Lipid breakdown 342

24 Vegetative growth of plants 34324.1 Introduction 34324.2 Composition of shoots 344

24.2.1 Plant cell walls 34424.3 Important shoot crops and their products 347

24.3.1 Temperate grasses 347. 24.3.2 Tropical grasses 348

xiv Contents

24.3.324.3.424.3.524.3.624.3.724.3.8

Forage legumesBrassicasStrawsHaySilageWood

24.4 Important root and tuber crops24.4.124.4.2

Root cropsTubers

348348348348349349351351351

25 Reproductive growth 35325.1 Flowering 35325.2 Fruit development and composition 35325.3 Fruit ripening 354

25.3.1 Changes in colour 35425.3.2 Changes in texture 35425.3.3 Changes in flavour 35525.3.4 Respiration in ripening fruit 355

25.4 Seed development 35625.4.1 Starch biosynthesis 35625.4.2 Protein synthesis 35725.4.3 Biosynthesis of fats 357

26 Plant nutrition 35926.1 Introduction 35926.2 Biochemical functions of major plant nutrients 359

26.2.1 Nitrogen 35926.2.2 Sulphur 35926.2.3 Phosphorus 36126.2.4 Potassium 36226.2.5 Calcium 36226.2.6 Magnesium 362

26.3 Trace elements - micronutrients 36226.4 Toxic effects of minerals 36226.5 Interaction between carbon and nitrogen metabolism 363

26.5.1 Carbon assimilation 36426.5.2 Nitrogen assimilation 36426.5.3 Senescence and nutrient cycling 366

27 Regulation of plant growth and development 36827.1 Introduction 36827.2 Responses to light 368

27.2.1 Effects on photosynthesis 368: 27.2.2 Phytochrome-mediated responses 368

27.2.3 Other responses to light 37027.3 Responses to temperature 370

27.3.1 Photosynthesis 37027.3.2 Vernalization 370

Contents xv

27.4 Responses to atmosphere 37027.5 Responses to stress 371

27.5.1 Temperature stress 37127.5.2 Water stress 37327.5.3 Salt stress 373

27.6 Nature of plant hormones 37427.7 Auxins 374

27.7.1 Biochemistry of auxins 37427.7.2 Synthetic auxins 37527.7.3 Sites of synthesis and transport of auxins 37627.7.4 Physiological activities and applications of auxins 377

27.8 Gibberellins 37927.8.1 Biochemistry of gibberellins 37927.8.2 Sites of synthesis and transport of gibberellins 37927.8.3 Physiological activities and applications of gibberellins 37927.8.4 Growth retardants 382

27.9 Cytokinins 38427.9.1 Biochemistry of cytokinins 38427.9.2 Sites of synthesis and transport of cytokinins 38427.9.3 Physiological activities and applications of cytokinins 384

27.10 Abscisic acid (ABA) 38727.10.1 Biochemistry of abscisic acid 38727.10.2 Sites of synthesis and transport of abscisic acid 38727.10.3 Physiological activities and applications of abscisic acid 387

27.11 Ethylene 38827.11.1 Biochemistry of ethylene 38827.11.2 Sites of synthesis and transport of ethylene 38927.11.3 Methods of modulating the effects of ethylene on plants 38927.11.4 Physiological activities and applications of ethylene 389

27.12 Miscellaneous'plant growth regulators 39227.12.1 Morphactins 39227.12.2 Maleic hydrazide 39227.12.3 Glyphosine 392

PART FOUR: STRATEGIES FOR PROCESSING OF NUTRIENTS IN ANIMALS 393

28 Digestion and absorption in ruminants and non-ruminants 39528.1 Introduction 39528.2 Structure of the digestive tract 39528.3 Carbohydrate digestion in monogastric animals 39628.4 Carbohydrate digestion in ruminants 39828.5 Absorption and utilization of glucose 39828.6 Digestion of lipids in monogastric animals 399

28.6.1 Digestion in the stomach 39928.6.2 Digestion in the small intestine 400

28.7 Absorption of lipid from the small intestine 40228.7.1 Glycerides and fatty acids 402

xvi Contents

28.7.2 Phospholipids 40328.7.3 Cholesterol 40328.7.4 Chylomicron formation 403

28.8 Uptake of absorbed lipid by body tissue 40328.9 Digestion of lipids in ruminant animals 40428.10 Lipid digestion in poultry 40628.11 Digestion of proteins in monogastric animals 406

28.11.1 Digestion in the stomach 40628.11.2 Digestion in the small intestine 407

28.12 Digestion of protein in ruminants 40928.13 Inhibitors of digestive enzymes 410

29 Maintenance 41329.1 Introduction 41329.2 Body temperature and heat production 41329.3 Biochemical production of heat 414

29.3.1 Background heat production from the maintenance of ion gradients 415in cells

29.3.2 Background heat production from protein turnover 41629.3.3 Background heat production from other metabolic events 41729.3.4 Heat production from muscular activity 41829.3.5 Heat production from uncoupled phosphorylation 419

30 Regulation and manipulation of growth and development in animals 42130.1 Introduction 421

30.1.1 General principles 42230.2 Rates and patterns of growth 42230.3 Muscle growth 424

30.3.1 Cellular growth as a component of muscular growth 42430.3.2 Protein accretion as a component of growth 42530.3.3 Control of protein synthesis 425

30.4 Growth of collagen 42530.5 Growth of bone 427

30.5.1 Calcification of bone 42830.6 Growth of adipose tissue 429

30.6.1 Growth in adipose tissue cell number 42930.6.2 Deposition of fat within adipocytes 430

30.7 Manipulation of growth 43130.7.1 Growth hormone and insulin-like growth factors 43130.7.2 Oestrogens and androgens 43330.7.3 The p-agonists 43530.7.4 Glucocorticoids 43930.7.5 Thyroid hormones 43930.7.6 Antibiotics 440

31 Lactation and its manipulation 447'31.1 Introduction 44731.2 Origins of the components of milk. 448

Contents xvii

31.3 The origin of lactose 44931.4 Milk proteins 451

31.4.1 The origin of milk proteins 45131.4.2 Amino acid supply to the mammary gland 45131.4.3 Protein synthesis 451

31.5 The fats 45231.5.1 Synthesis de novo 45231.5.2 Uptake of fatty acids from blood 45331.5.3 Modifications to fatty acids in the mammary gland 453

31.6 The supply of energy in the mammary gland 45431.7 Metabolism in lactation. 454

31.7.1 Endocrine control of lactation 45431.8 Manipulation of lactation 455

31.8.1 Dietary manipulation of lactation 45631.8.2 Manipulation of milk production by exogenous hormones 457

32 Muscle and meat 45932.1 Introduction 45932.2 Biochemistry of muscular contraction 459

32.2.1 Structure of thick filaments 46132.2.2 Structure of thin filaments 46132.2.3 Mechanism of muscle movement 46132.2.4 Control of muscle movement 462

32.3 Energy provision in muscle tissue 46432.3.1 Myoglobin 465

32.4 Changes in muscle after death 46532.4.1 Enzymes leading to the degeneration of myofibrils 46632.4.2 The role of myoglobin 46632.4.3 Conditioning 46732.4.4 Cold shortening 46732.4.5 Effects of stress pre-slaughter 467