Chapter 3: Amino Acids, Peptides, and Proteins

1. Chapter 3: Amino Acids, Peptides, and Proteins Dr. Rita P.-Y. Chen (陳佩燁) Assistant Research Fellow Institute of Biological Chemistry Academia Sinica

2. Amino acid • Chiral center • Optically active: rotate plane-polarized light • Amino acid in protein: L stereoisomer

3. Absolute configuration: D,L system Not all L-amino acids are levorotatory (rotating polarized light to the left)

4. Specifying onfiguration: RS system L-amino acid has S configuration

9. aliphatic amino acid structures CH3 CH Isoleucine (I) (Ile) CH2 CH3

10. Side Chains with Alcohol Groups • Serine (Ser, S) and Threonine (Thr, T) have uncharged polar side chains Catalytic role, phosphorylation, o-linked glycosylation, hydrogen bond

11. Structures of aspartate, glutamate, asparagine and glutamine

12. Structures of histidine, lysine and arginine d-Guanido group imidazole group

13. Methionine and cysteine First a.a. Catalytic role, disulfide bond

14. Fig 3.4 Formation of cystine Disulfide bond

15. Aromatic amino acid structures Indole group phosphorylation

16. Proline has a nitrogen in the aliphatic ring system • Proline (Pro, P) - has a three carbon side chain bonded to the a-amino nitrogen • The heterocyclic pyrrolidine ring restricts the geometry of polypeptides

20. p t

24. Uncommon a.a. Prothrombin Ca2+ binding protein Plant cell wall Collagen Collagen elastin myosin 21th a.a. Added during protein synthesis UGA codon glutathione peroxidases

25. Other amino acids (not constituents of proteins) : metabolite Key intermediate in biosynthesis of Arg and in urea cycle Urea p843

26. 補充教材

27. Amino acid can act as acids and bases

28. 較易解離 較易解離

29. The characteristic pH at which the net electric charge is zero is called the isoelectric point or isoelectric pH, designated pI. For glycine, which has no ionizable group in its side chain, the isoelectric point is simply the arithmetic mean of the two pKa values:

30. 0 pI = (2.19+4.25)/2 = 3.22

31. 0 pI = (6+9.17)/2 = 7.59

32. Peptides and Proteins

33. SGYAL

34. 9.6 -1 +2 +1 0 4.25 pI = (4.25+9.6)/2 =6.93 10.53 2.34

35. Artificial Sweetener, ex. in Diet Coke • 200 times sweeter than sugar • D-form a.a. substitution is bitter • 苯酮尿症(Phenylketonuria) 患者不可使用, use Alatame instead

36. 催產素 血管收縮素 Fig. 3-13, p.75

37. Glutathione • an important water-phase antioxidant and essential cofactor for antioxidant enzymes

38. Protein size is varied

39. Polypeptides have characteristic amino acid compositions

40. Some proteins have chemical groups other than a.a. • Non a.a. part – prosthetic group

42. Protein purification • Fractionation: protein solubility depends on temperature, pH, salt • Dialysis • Ultrafiltration: N2 purge, centrifugation • Column chromatography • Electrophoresis

43. Column chromatography

44. Cation exchange chromatography: • Protein carries postive charge (cation) • Buffer pH must be lower than protein pI • No sample volume limit

45. Size exclusion chromatography • Big protein runs faster • Sample volume is limited • Column is usually long

46. Affinity chromatography • Separate proteins by their binding specificities • No sample volume limit

48. Electrophoresis: (1) SDS-PAGE • Proteins migrate according to their size and shape • One SDS bind for every two residues • Protein is denatured, subunits will be separated cathode Anode

49. Coomassive blue staining 4 subunits