Amino Acid Biosynthesis

1. Amino Acid Biosynthesis Introduction

2. Essential (10) Arginine Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine Non-essential Alanine Asparagine Aspartate Cysteine Glutamate Glutamine Glycine Proline Serine Tyrosine Essential and Non-essential Amino Acids

3. Importance of Balanced Diet • Milk: proper balance of amino acids • Beans: lysine rich; methionine poor • Wheat: methionine rich; lysine poor • Bioengineered Wheat: high lysine content

4. Nitrogen Acquisition • Nitrogen Fixation • Nitrate Assimilation • Ammonium Assimilation

5. Nitrogen Fixation(few strains of bacteria)

6. Nitrifying Bacteria

7. Nitrate Assimilation(Green plants, some fungi and bacteria)

8. Ammonium Assimilation (1)(Carbamyl Phosphate Synthetase)

9. Ammonium Assimilation (2)(Biosynthetic Glutamate Dehydrogenase)and/or(Glutamine Synthetase)

10. Microbial Nitrogen Acquisition(Metabolic Sources of Organic Nitrogen)

11. Microbial Ammonium Assimilation(High NH3)

12. Mechanism of Glutamine Synthetase

13. Microbial Ammonium Assimilation(Low NH3)

14. Microbial Ammonium Assimilation • High [NH3] • “Biosynthetic” Glutamate Dehydrogenase • Glutamine Synthetase • Low [NH3] • Glutamine Synthetase • Glutamate Synthase (Glutamate Oxoglutarate Aminotransferase, GOGAT)

15. Microbial Ammonium Assimilation(High NH3)

16. Microbial Ammonium Assimilation(Low NH3)

17. Role of Glutamate(Nitrogen Donor)

18. Role of Glutamine(Nitrogen Donor)

19. Specific Products of Glutamine • Histidine • Tryptophan • Carbamyl Phosphate (CPSII) • Glucosamine • ATP • CTP

20. Regulation of Glutamine Synthetase

21. Pathways of Nitrogen Incorporation

22. Products Histidine Tryptophan Carbamyl-P Glucosamine ATP CTP Nitrogen Status Alanine Serine Glycin Feedback Inhibition(Nitrogen-sufficient conditions)

23. Modulation of Enzyme ActivitySensitivity to [NH3] Importance of Glutamine/a-ketoglutarate Ratio

24. Control of Enzyme Activity(Escherichia coli Enzyme)

25. Control of Gene Expression GlnA = Glutamine Synthetase NtrB = Autokinase NtrC = Transcription Factor

26. Control of Gene Expression(NtrC Transcription Factor)

27. Nitrogen Sufficiency (Rich)(Low Expression of Glutamine Synthesis) • Basal expression from P1 (weak promoter) • Termination at rho-independent terminator • Basal expression from Pinternal • RNA polymerase (s70)

28. Control of Gene Expression(Nitrogen Sufficiency)

29. Nitrogen Limitation(Ammonia Assimilation = High Expression) • Expression from P2 (strong promoter) • Requires NtrC–P (binds to enhancer) • RNA polymerase (s54) • Reads through terminator • Upregulation of ntrB and ntrC

30. Control of Gene Expression(Nitrogen Limitation)

31. Two Component Sensor-Response Regulator System NtrB and NtrC

32. Two Component Sensor-Response Regulator Systems • Autokinase Sensor Proteins (NtrB) (autophosphorylation of histidine residue) • Sensor domain • Transmitter domain (C-terminus) • Response Regulators (NtrC) • N-terminal receiver domain • Cross-regulation

33. NtrC

34. Other NtrC-Sensitive Operons • Direct effect of NtrC-P (e.g. hut operon) • Indirect effect of NtrC-P

35. Conclusion Complex regulation of glutamine synthetase confirms important role in nitrogen metabolism

36. Biosynthesis of Non-essential Amino Acids Simple Pathways

37. Alanine

38. Aspartate and Asparagine

39. Glutamate and Glutamine

40. Proline

41. Proline (continued)

42. Arginine(N-Acetylglutamate Synthase)

43. Arginine (continued)(Acetylglutamate Kinase)

44. Arginine (continued)(N-Acetyl-g-glutamyl-P Reductase)

45. Arginine (continued)(N-Acetylornithine g-Aminotransferase)

46. Arginine (continued)(Acetylornithine Deacetylase)

47. Arginine (continued)(Acetylornithine:Glutamate Acetyltransferase)

48. Arginine (continued)(Ornithine Transcarbamylase)

49. Arginine (continued)(Argininosuccinate Synthetase)

50. Arginine (continued)(Argininosuccinase)