1. Quiz 2: Definitions Monosaccharide Disaccharide Oligosaccharide Polysaccharide Aldose Ketose Lipid Fatty Acid Amphiphillic Amphipathic Saturated Unsaturated

2. Quiz 2: Identification (Carbs)

3. Quiz 2: Identification (Lipids) #Cs Common Name IUPAC Name 12Lauric AcidDodecanoic Acid 14Myristic AcidTetradecanoic Acid 16Palmitic AcidHexadecanoic Acid 18Stearic AcidOctadecanoic Acid 18Oleic Acid9-Octadecenoic Acid 18Linoleic Acid 9,12-Octadecadienoic Acid 18  -Linoleic Acid 9,12,15-Octadecatrienoic Acid

4. Protein Structure III

5. Relevent Interactions Hydrophobic Forces Electrostatic forces –Ion Pairs –Dipole–Dipole Interactions –Hydrogen Bonding Covalent Bonds

6. Hydrophobic Forces (Entropic) Minimizes order of solvent H 2 O which occurs when hydrophobic molecules are in aqueous environment Primary Determinant of Tertiary and Quaternary Structures

7. Hydrophobic Interactions Not “bonds”; exclusion of water Buried in interior Important in tertiary structure

8. Electrostatic Forces Ion Pairs Attractive or Repulsive Coulomb’s Law (strength proportional to 1/r 2 Competition between buried ionic interactions and hydrated ionic species on the surface

9. Ion Pairs Strength dependent on magnitude of charges, dielectric constant, and distance Modest strength Contribute little to native protein structure

10. Figure 6-36 Ion Pairs or Salt Bridges (Myoglobin)

11. Ion-Polar Bonds Similar to electrostatic bonds Alternative to interactions with water Contribute little to native protein structure

12. Dipole-Dipole Interactions Van der Waals Attractions Strength proportional to 1/r 6 Dipole–Dipole Interactions Dipole-Induced Dipole Interactions Induced Dipole-Induced Dipole Interactions (London Dispersion Forces) Significant Contribution to Protein Native Structure

13. Dipole-Dipole Interactions

14. Typical Hydrogen Bonds between side chains (unshared electron pair: N and O)

15. Hydrogen Bonds Strength greatest in a polar environment Contribute greatly to secondary structure

16. Other Hydrogen Bonds (e.g. –SH) Similar electronegativity as –CH 3 More polarizable than –CH

17. Covalent Bonds Disulfide Bonds

18. Covalent (Disulfide) Bonds Do not need to be adjacent in primary structure Strong Intra- or Interchain

19. Sum of Forces Conformational Stability e.g. linking of fingers

20. Flexibility (conformational changes possible) Interaction (binding) of small molecules (effectors) Modification of protein amino acids – e.g. phosphorylation of serine

21. Figure 6-37 Cys 2 –His 2 Zinc Finger Motif Stabilization of Small Domain

22. Figure 6-38 Molecular Dynamics of Myoglobin Proteins Are Dynamic Structures

23. Tertiary Structure Folding and ordering of a polypeptide chain due to interactions involving the amino acid side chains

24. Characteristics of Tertiary Structure May contain both  helices and  sheets Structural characteristics –Nonpolar residues: interior –Charged residues: surface (hydrated) –Polar residues: surface (hydrated) or interior (hydrogen-bonded) –Compact: little or no internal space for water molecules Domain Structure

25. Figure 6-27 Side Chain Distribution in Horse Heart Cytochrome c

26. Tertiary Structures Contain Combinations of Secondary Structure Motifs

27. Figure 6-28 Super Secondary Structural Motifs   hairpin  Greek Key

28. All  -Helix Proteins

29. All  -Sheet Proteins

30.  Proteins

31. Large Proteins Form Domains

32. Figure 6-31 Two Domain Protein (glyceraldehyde-3-P dehydrogenase)

33. Quaternary Structure Specific association of polypeptide chains Subunits

34. Characteristics of Quaternary Structure Identical or nonidentical subunits Subunits usually associate noncovalently Subunits are symmetrically arranged Efficient means of producing highly complex proteins Basis for regulatory behavior of many enzymes

35. Figure 6-33 Quaternary Structure of Hemoglobin (  2  2 )

36. Three Broad Categories of Proteins Fibrous Proteins Globular Proteins Membrane Proteins

37. Characteristics of Fibrous Proteins Rod-like Insoluble –due to hydrophobic AAs both inside and outside Structural

38.  -Keratin: Evolved for Strength (Hair, Wool, Nails) Right handed  -helix Left handed coiled coil

39. Figure 6-15a Coiled Coil non-polar residues

40.  -Keratins - Crosslinked by Disulfide Bonds

41. Collagen — A Triple Helical Cable Component of connective tissue Distinctive amino acid compostion –~33% glycine –15-30% 4-hydroxyproline –Some 3-hydroxyproline & 5-hydroxylysine Right-handed triple helix Organized into fibrils Fibrils are covalently cross-linked Collagen defects are responsible for a variety of human diseases

42. Collagen: in connective tissue, cartilage, gelatin left-handed  -helix 3 AA per turn 3  -chains are supertwisted mainly Gly, Ala, Pro (Gly-X-Y motif) 6% hydroxy-proline confers thermostability

43. Proline Hydroxylase Requires ascorbic acid (vitamin C) Scurvy

44. Globular Proteins

45. Properties of Globular Proteins Majority of proteins –Dynamic Functions, i.e. enzymes Mixture of secondary structures Soluble –hydrophobic core, polar surface

47. Water (aqueous) Non-Polar Membrane Proteins: receptors, transporters, enzymes, ion channels Hydrophobic

48. Determination of Tertiary Structure X-Ray Crystallography Nuclear Magnetic Resonance (NMR)

49. Protein Crystals

50. Figure 6-21 X-Ray Diffraction Pattern

51. Figure 6-22 Thin Section Electron Density Map

52. Solving the first protein crystal structure (1958 Dickerson, RE; A little ancient history; Protein science, 1992

53. Figure 6-23 Electron Density Map Resolution (Diketopiperazine)

54. Structure determination of molecules in solution shows conformational changes/heterogeneity of molecules in solution size limit (< 30,000 Da) Nuclear Magnetic Resonance (NMR)

55. Figure 6-24 Nuclear Overhauser Spectroscopy (NOESY) Spectrum of a Protein

56. 43 Different Conformations of Brazzein in Solution

57. Protein Stability (largely through many weak non-covalent interactions) “Proteins are only marginally stable entities under physiological conditions” “A protein structure is the result of a delicate balance among powerful countervailing forces”

58. Protein Folding Anfinsen’s classic experiment How proteins find their native structure ribosome mRNA nascent polypeptide folding Folded polypeptide

59. Page 159 Chaotropic Agents

60. Unfolding Denaturation Refolding Renaturation

61. What Drives Protein Folding? Unfolded state: high entropy, weak interactions between AA side-chains and water Folded state: many weak non-covalent interactions (H-bonds, salt bridges, van der Waals) and covalent interactions (disulfide bonds) Protein tries to reach the thermodynamically most stable state! Protein conformation with lowest free energy is the one with the maximum number of weak interactions.  G folding : -(20-65) kJ/mol

62. How Do Proteins Fold? Levinthal Paradox: It CANNOT be a random search 1 AA:10 different conformations (backbone + side-chains) 100 AA:10 100 different conformations 1 conformation tried within 10 -13 s (shortest possible time) 10 100 conformations: 10 77 years!!!

63. Protein Folding Pathway Secondary structure (ms-s) Hydrophobic collapse (ms) Tertiary structure elements (s)

64. Folding Funnels

65. In Vitro vs. in Vivo Folding in vivo, protein folding is usually highly efficient (E. coli: 100 AA protein / 5 s) in vitro, protein folding is often problematic and very inefficient biggest problem: Protein aggregation In vivo: Molecular Chaperones

66. Molecular Chaperones proteins that assist other proteins in their folding to the native state prevent non-productive side reactions such as irreversible aggregation do not form part of the final structure do not contain information about the folding pathway

67. Molecular Chaperones Prevent Protein Aggregation U nfolded U1U1 aggregates nascent polypeptide + chaperone molecular chaperone F olded

68. Quaternary Structure of the GroE Chaperone

69. Figure 6-42 Mechanism of Protein Disulfide Isomerase

70. Table 6-4 Protein Misfolding Diseases