The document discusses various aspects of peptide structure and function including: 1. The common secondary structural motifs of peptides including alpha helices, beta sheets, beta turns, and random coils. 2. Techniques used to study peptide conformation such as crystallography, NMR, and CD spectroscopy. 3. Factors that influence peptide structure including amino acid sequence, solvent environment, and intermolecular interactions.

1. PEPTIDE STRUCTURE - FUNCTION

2. Rational Design of Peptides - Driving Force CRYSTALLOGRAPHY NMR – HIGH RES ABS, FLUORES, CD, IR - LOW SEQUENCING SEQUENCE STRUCTURE

3. + PEPTIDE BOND FORMATION AA1 AA2 H 2 0 DIPEPTIDE PROTEASES H 2 NC  HC OH O R HHNC  HC OH O R H 2 NC  HC O R 1 HNC  HC OH O R 2

4. PHYSICO-CHEMICAL PROPERTIES PHYSICAL PROPERTIES ADDITIVE LEGNTH, MASS NOT ADDI IVE Pka => AA1+AA2 ===== DIPEPEPTIDE ENERGETICS, REACTIVITY ETC

5. STRUCTURE OF THE PEPTIDE BOND

6. GEOMETRICAL CONSTRAINTS - CONFORMATIONS ALLOWED NOT-ALLOWED ANGLES

7. DIPOLE ORIGIN OF PEPTIDE BOND PLANE NOT ALL CONFORMATIONS POSSIBLE PREFERED CONFORMATIONS

8. STRUCTURAL MOTIFS - FUNCTIONAL HELIX -  -HELIX, 3-10 HELIX  -SHEETS (Parallel, Anti-Parallel  -TURNS RANDOM COILS

9.  helix α-helix (30-35%) Hydrogen bond between C=O (carbonyl) & NH (amine) groups within strand (4 positions apart) 3.6 residues / turn, 1.5 Å rise / residue Typically right hand turn Most abundant secondary structure α-helix formers: A,R,L,M,E,Q,K

10. the alpha-helix: repeating i,i+4 h-bonds 2 1 3 4 5 7 8 9 6 10 11 12 right-handed helical region of phi-psi space hydrogen bond

11. The  -helix, with i,i+4 h-bonds, is not the only way to have local hydrogen bonding of the backbone to itself. The 3 10 helix has hydrogen bonds between residues i and i+3 The  helix has hydrogen bonds between residues i and i+5. For a number of reasons almost all helices in proteins are  -helices--include backbone, side chain steric issues, van der Waals contacts, H-bond geometry  -helix 3 10 helix  helix these are poly-Ala, so the gray balls on the outside are  -carbons from the side chains

12.  sheet &  turn β-sheet / β-strand (20-25%) Hydrogen bond between groups across strands Forms parallel and antiparallel pleated sheets Amino acids less compact – 3.5 Å between adjacent residues Residues alternate above and below β-sheet β-sheet formers: V,I,P,T,W

14. β-turn Short turn (4 residues) Hydrogen bond between C=O & NH groups within strand (3 positions apart) Usually polar, found near surface β-turn formers: S,D,N,P,R TURNS

15. Others Loop (bridging region) Regions between α-helices and β-sheets On the surface, vary in length and 3D configurations Do not have regular periodic structures Loop formers: small polar residues Coil (40-50%) Generally speaking, anything besides α-helix, β-sheet, β-turn

16. Principal types of secondary structure found in proteins Repeating (  ) values -63 o -42 o -57 o -30 o -119 o +113 o -139 o +135 o   -helix (1  5) (right-handed)    helix (1  4) Parallel  -sheet Antiparallel  -sheet

17. STRUCTURES IN ACTION GCN4 “leucine zipper” (green) bound as a dimer (two copies of the polypeptide) to target DNA The GCN4 dimer is formed through hydrophobic interactions between leucines (red) in the two polypeptide chains Leu Leu

18. TECHNIQUES – PEPTIDE COMFORMATION CD X-ray Crystallography NMR

19. Do Small Peptides have Conformation Yes & No. S-Peptide Ribonuclease A – Helical structure in solution Use of Helix Inducing Solvents – TFE and N-Propanol

20. Helix Induction and Propensity

21. CONFORMATIONAL TRANSITION

23. PROPENSITY CALCULATION

24. BIOLOGY OF PEPTIDES RIBOSOMAL PROTEINS NON-RIBOSOMAL PEPTIDES SPECIFIC ENZYMES PROTEOLYTICALY PROCESSED DEGRADED TO AA (Antibiotics, phytochelatins) (Enzymes) MHC Peptides

25. CHEMICAL METHOD –PEPTIDE SYNTHESIS STAGE 1: ASSEMBLE AA ON POLYMER SUPPORT (R – PROTECTED) NON-REACTIVE STAGE 2: CLEAVE THE SYNTHESIZED PEPTIDE a) CLEAVAGE OF CHAIN b) DE-PROTECT SIDE CHAIN STAGE 3: PURIFY CRUDE PEPTIDES – HPLC STAGE 4: STORAGE – LYOPHILIZE, SPEEDVAC, ETC STAGE 5: SEQUENCE, MALDI-TOF

26. SOLID-PHASE PEPTIDE SYNTHESIS (SPPS) STAGE 1: a) Attach N-terminal + Side Chain Protected to Polymer Support (Activation of C & Coupling to Support) b) Deprotection (N-term ) c) Coupling Next AA (Protected) d) Deprotection (N-term) Continued ….

27. V 8 Protease “ Conformational Trap” Protease-mediated Protein Splicing – Nature’s Choice LYGSTSQE VASVKQAFDAVGVK NH-VASVKQAFDAVGVK-OH NH-LYGSTSQE-OH “ Proteolysis” “ Reverse Proteolysis” LYGSTSQE VASVKQAFDAVGVK

28. TAAAKFE “ Conformational Trap” can act alone

29. Conformational Trap of product – ambient conditions, easy Isolation, and Purification of Products Applications 1. Ability to Incorporate Non-Natural aminoacids or synthesize Man-made peptides or proteins of therapeutic interest Semisynthetic Insulin, Hemoglobin, and IL-10 Sortases – Glycoprotein synthesis Laboratory reagents :- Protein with reporter groups, Kinases or Phosphotases with Pmp(phosphonomethylene phenylalanine )