1. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 1 Chapter 10 Metabolism: The Use of Energy in Biosynthesis

2. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 2 Anabolism from a carbon source and inorganic molecules, microbes synthesize new organelles and cells –a lot of energy is required for biosynthesis turnover –continual degradation and resynthesis of cellular constituents by nongrowing cells metabolism is carefully regulated –for rate of turnover to be balanced by rate of biosynthesis –in response to organism’s environment

3. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 3 Figure 10.1

4. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 4 Table 10.1

5. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 5 Principles Governing Biosynthesis 1.macromolecules are synthesized from limited number of simple structural units (monomers) –saves genetic storage capacity, biosynthetic raw material, and energy 2.many enzymes used for both catabolic and anabolic processes –saves materials and energy

6. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 6 More principles… 3. catabolic and anabolic pathways are not identical, despite sharing many enzymes –permits independent regulation Figure 10.2

7. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 7 More principles… 4. to synthesize molecules efficiently, anabolic pathways must operate irreversibly in the direction of biosynthesis –done by coupling breakdown of ATP to certain reactions in biosynthetic pathways –drives the biosynthetic reaction to completion 5. in eucaryotes, anabolic and catabolic reactions located in separate compartments –allows pathways to operate simultaneously but independently

8. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 8 More principles… 6. catabolic and anabolic pathways use different cofactors –catabolism produces NADH –NADPH used as electron donor for anabolism large assemblies (e.g., ribosomes) form spontaneously from macromolecules by self-assembly

9. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 9 Precursor Metabolites generation of precursor metabolites is critical step in anabolism carbon skeletons are used as starting substrates for biosynthetic pathways –examples are intermediates of the central metabolic pathways –most are used for the biosynthesis of amino acids

10. The importance of CO2 in macromolecule biosynthesis 1. CO2 is taken in anabolism via fixation step during carboxylation phase. 2. CO2 later is converted to organic carbon. 3. The organic carbon generates the precursor metabolites as it generates ATP. 4. Precursor metabolites are carbon skeletons used as the starting substrates for the synthesis of monomers and other building blocks. 5. Monomers needed for the synthesis of macromolecules. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 10

11. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 11 The Organization of Anabolism Figure 10.3

12. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 12 The Fixation of CO 2 by Autotrophs- important the Calvin cycle the reductive TCA cycle the acetyl-CoA pathway the hydroxypropionate cycle

13. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 13 Calvin cycle used by most autotrophs to fix CO 2 also called the reductive pentose phosphate cycle in eucaryotes, occurs in stroma of chloroplasts in cyanobacteria, some nitrifying bacteria, and thiobacilli, may occur in carboxysomes –inclusion bodies that may be the site of CO 2 fixation

14. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 14 Calvin Cycle consists of 3 phases –the carboxylation phase –the reduction phase –the regeneration phase three ATPs and two NADPHs are used during the incorporation of one CO 2

15. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 15 Figure 10.4

16. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 16 Calvin Cycle the carboxylation phase –catalyzed by the enzyme ribulose 1.5- bisphosphate carboxylase, also called ribulose bisphosphate carboxylase/oxygenase (rubisco) –rubisco catalyzes addition of CO 2 to ribulose-1,5-bisphosphate (RuBP), forming 2 molecules of 3- phosphoglycerate

17. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 17 The Carboxylation Phase Figure 10.4 catalyzed by the enzyme ribulose 1.5-bisphosphate carboxylase, also called ribulose bisphosphate carboxylase/oxygenase (rubisco) rubisco catalyzes addition of CO 2 to ribulose-1,5- bisphosphate (RuBP), forming 2 molecules of 3- phosphoglycerate

18. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 18 The Ribulose 1.5-Bisphosphate Carboxylase Reaction Figure 10.5

19. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 19 The Reduction Phase 3-phospho- glycerate reduced to glyceraldehyde 3-phosphate Figure 10.4

20. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 20 The Regeneration Phase RuBP regenerated carbohydrates (e.g., fructose and glucose) are produced Figure 10.4

21. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 21 Summary 6CO 2 + 18ATP + 12NADPH + 12H + + 12H 2 O  glucose + 18ADP + 18P i + 12NADP +

22. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 22 Other CO 2 -Fixation Pathways the reductive TCA cycle –used by some chemolithoautotrophs –runs in reverse direction of the oxidative TCA cycle

23. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 23 Figure 10.6

24. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 24 Other CO 2 -Fixation Pathways the hydroxypropionate cycle –used by some archael genera and the green nonsulfur bacteria (also anoxygenic phototrophs)

25. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 25 Figure 10.7

26. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 26 Other CO 2 -Fixation Pathways the acetyl-CoA pathway –methanogens use portions of the acetyl- CoA pathway for carbon fixation –involves the activity of a number of unusual enzymes and coenzymes

27. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 27 Figure 10.8: The Acetyl-CoA Pathway

28. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 28 Synthesis of Sugars and Polysaccharides gluconeogenesis –used to synthesize glucose and fructose from noncarbohydrate precursors

29. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 29 Gluconeogenesis synthesis of glucose and related sugars from nonglucose precursors –glucose, fructose and mannose are gluconeogenic intermediates or made directly from them –galactose is synthesized with nucleoside diphosphate derivatives –bacteria and algae synthesize glycogen and starch from adenosine diphosphate glucose

30. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 30 Gluconeogenesis functional reversal of glycolysis, but the two pathways are not identical –7 enzymes shared –4 enzymes are unique to gluconeogenesis

31. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 31 Figure 10.9

32. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 32 Synthesis of Monosaccharides several sugars are synthesized while attached to a nucleoside diphosphate such as uridine diphosphate glucose (UDPG) Figure 10.10

33. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 33 Uridine Diphosphate Galactose and Glucuronate Synthesis Figure 10.11

34. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 34 Synthesis of polysaccharides also involves nucleoside diphosphate sugars –e.g., starch and glycogen synthesis ATP + glucose 1-P  ADP-glucose + PP i (glucose) n + ADP-glucose  (glucose) n+1 + ADP

35. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 35 Peptidoglycan Synthesis complex process involves use of UDP derivatives also uses bactroprenol, a lipid carrier, to transport NAG-NAM-pentapeptide units across the cell membrane cross links are formed by transpeptidation

36. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 36 Figure 10.12

37. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 37 Bactoprenol is attached to N-acetylmuramic acid (NAM) Figure 10.13

38. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 38 Figure 10.14

39. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 39 Patterns of Cell Wall Formation autolysins –carry out limited digestion of peptidoglycan –activity allows new material to be added to wall 2 general patterns of cell wall formation

40. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 40 growth at one or just a few sites usually at site of septum formation observed in many gram-positive bacteria growth sites scattered along length of cell growth also at site of septum formation observed in many rod-shaped bacteria Figure 10.15

41. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 41 The Synthesis of Amino Acids precursor metabolites used as starting substrates for synthesis of amino acids –carbon skeleton is remodeled –amino group and sometimes sulfur are added

42. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 42 The Synthesis of Amino Acids… nitrogen addition to carbon skeleton is an important step –potential sources of nitrogen: ammonia, nitrate, or nitrogen most cells use ammonia or nitrate –ammonia nitrogen easily incorporated into organic material because it is more reduced than other forms of inorganic nitrogen

43. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 43 Ammonia Incorporation into Carbon Skeletons –two mechanisms reductive amination –ammonia N can be directly assimilated by transaminase activity or the glutamate dehydrogenase or the glutamine synthetase-glutamate synthase systems

44. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 44 Ammonia Incorporation into Carbon Skeletons two mechanisms –reductive amination –glutamine synthetase-glutamate synthase systems once incorporated, nitrogen can be transferred to other carbon skeletons by transaminases

45. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 45 Ammonia Assimilation Pathway Figure 10.16

46. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 46 Glutamine Synthetase and Glutamate Synthase (GOGAT) System Figure 10.17

47. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 47 Ammonia Incorporation using Glutamine Synthetase and Glutamate Synthase Figure 10.18

48. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 48 Assimilatory Nitrate Reduction used by bacteria to reduce nitrate to ammonia and then incorporate it into an organic form nitrate reduction to nitrite catalyzed by nitrate reductase reduction of nitrite to ammonia catalyzed by nitrite reductase Figure 10.19

49. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 49 Nitrogen Fixation reduction of atmospheric nitrogen to ammonia catalyzed by nitrogenase –found only in a few species of procaryotes requires large ATP expenditure Figure 10.20

50. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 50 Figure 10.21

51. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 51 Mechanism of nitrogenase activity repeated 3 times to reduce N2 to 2 molecules of ammonia Figure 10.22

52. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 52 Sulfur Requirements of Cell sulfur needed for –synthesis of amino acids cysteine and methionine –synthesis of several coenzymes sulfur obtained from –external sources –intracellular amino acid reserves

53. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 53 Use of Sulfate as a Sulfur Source sulfate = inorganic sulfur source –assimilatory sulfate reduction sulfate reduced to H 2 S and then used to synthesize cysteine

54. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 54 Assimilatory Sulfate Reduction involves sulfate activation through formation of phosphoadenosine 5’- phosphosulfate (PAPS) followed by reduction of the sulfate

55. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 55 Assimilatory sulfate reduction Figure 10.24 Figure 10.23 different than dissimilatory sulfate reduction, where sulfate acts as electron acceptor for anaerobic respiration activated sulfate (PAPS)

56. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 56 Assimilatory sulfate reduction Figure 10.23 different than dissimilatory sulfate reduction, where sulfate acts as electron acceptor for anaerobic respiration activated sulfate (PAPS) Figure 10.24

57. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 57 Formation of cysteine two processes used 1)H 2 S + serine  cysteine + H 2 O 2)serine + acetyl-CoA  O-acetylserine + Co-A O-acetylserine + H 2 S  acetate + cysteine

58. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 58 Amino Acid Biosynthesis – Branching Pathways used in the synthesis of multiple amino acids a single precursor metabolite can give rise to several amino acids biosynthetic pathways for aromatic amino acids also share intermediates

59. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 59 Sample pathways Figure 10.25Figure 10.26

60. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 60 Anaplerotic Reactions TCA cycle intermediates are used in many amino acid biosynthetic pathways replenishment of these intermediates is provided by anaplerotic reactions allow TCA cycle to function during periods of active biosynthesis e.g., anaplerotic CO 2 fixation e.g., glyoxylate cycle

61. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 61 Anaplerotic CO 2 fixation phosphoenolpyruvate (PEP) carboxylase –phosphoenolpyruvate + CO 2  oxaloacetate + P i pyruvate carboxylase –pyruvate + CO 2 + ATP + H 2 O  oxaloacetate + ADP + P i reaction requires the cofactor biotin

62. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 62 Glyoxalate cycle other anaplerotic reactions are part of the glyoxalate cycle, a modified TCA cycle Figure 10.27

63. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 63 The Synthesis of Purines, Pyrimidines, and Nucleotides most microbes can synthesize their own purines and pyrimidines purines –cyclic nitrogenous bases consisting of 2 joined rings –adenine and guanine pyrimidines –cyclic nitrogenous bases consisting of single ring –uracil, cytosine, and thymine nucleoside = nitrogenase base-pentose sugar nucleotide = nucleoside-phosphate

64. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 64 Phosphorus Assimilation phosphorus found in nucleic acids as well as proteins, phospholipids, ATP and some coenzymes most common phosphorus sources are inorganic phosphate and organic phosphate esters

65. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 65 Phosphorus Assimilation inorganic phosphate (P i ) –incorporated through the formation of ATP by: photophosphorylation oxidative phosphorylation substrate-level phosphorylation organic phosphate esters –present in environment in dissolved or particulate form –hydrolyzed by phosphatases, releasing P i

66. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 66 Purine Biosynthesis complex pathway in which several different molecules contribute parts to the final purine skeleton Figure 10.28 inosinic acid

67. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 67 Figure 10.29 deoxyribonucleotides formed by reduction of nucleoside diphosphates or nucleoside triphosphates initial products are ribonucleotides

68. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 68 Pyrimidine Biosynthesis ribonucleotides are initial products deoxy forms of U and C nucleotides formed by reduction of ribose to deoxyribose Figure 10. 30

69. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 69 Figure 10.31

70. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 70 Lipid Synthesis lipids –found in cell membranes –most contain fatty acids fatty acids –synthesized then added to other molecules to form other lipids such as triacylglycerols and phospholipids

71. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 71 Fatty Acids synthesized from acetyl-CoA, malonyl-CoA, and NADPH by fatty acid synthase system during synthesis the intermediates are attached to the acyl carrier protein double bonds can be added in two different ways

72. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 72 Fatty acid synthesis catalyzed by fatty acid synthetase involves activity of acyl carrier protein (ACP) Figure 10.32

73. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 73 Triacylglycerols and phospholipids made from fatty acids and glycerol phosphate phosphatidic acid is an important intermediate in this pathway

74. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 74 Triacylglycerols and phospholipids Figure 10.33

75. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 75 Phospholipids major components of eucaryotic and bacterial cell membranes synthesized from phosphatidic acid by forming CDP-diacylglycerol, then adding an amino acid