1. Secondary structure elements  helices  strands/sheets/barrels  turns The type of 2° structure is determined by the amino acid sequence –Chemical & physical characteristics –How? Area of research

2. Amino acid ‘flexibility’ Side chain interactions

3.  -turn Proteins  up to 1/3 turns and loops Common linker for  -sheets and  - helices 180 o turn involving 4 residues –H-bond between C=O and N-H Which AA?

4.  -turn Proline –Imino N  cis conformation (6%) Glycine –Very flexible Often found on the exterior of the folded protein: solvent exposed

5. 3 ° and 4 ° structure 3 ° structure –Overall 3-D arrangement –Interaction of 2° structural elements 4 ° structure –Arrangement of separate chains/subunits –Non-covalently linked Possible exception: disulfide bonds 2 classes of proteins –Fibrous proteins (extended) –Globular proteins (~spherical)

6. Fibrous proteins Structural roles Typically single type of 2 ° structure –Long strands of  helices (eg.  -keratin/collagen) –Big sheets of  structure (eg. silk) Insoluble in H 2 O –  conc of H-phobic on interior and surface –Buried by packing chains together Strong and flexible –(eg. hair, silk, cartilage)

7. Collagen Major constituent of connective tissues (bone, tendon, ligaments, skin…) Helical 2° structure distinct from  helix –3 AA/turn (tighter  –Left-handed (opposite twist) collagen “triple helix”  tropocollagen –Helix: 2° structure –Triple helix: 4° structure

8. Collagen Gly (35%), Ala (11%) and Pro (or HyPro) (21%) Every 3 rd residue is a Gly (Gly–X-Y-Gly–X-Y) –Genetic defects when G is changed (“mutated”) eg. osteogenesis imperfecta Chains linked by H-bonds –Backbone NH of Gly and backbone C=O of X in another chain Chains also linked by uncommon covalent bonds –Side chain linkage

9. Collagen Triple helix aligns and crosslinks  collagen fibrils –Crosslinked via covalent bonds between Lys, HyLys and His Too many crosslinks? –↓ flexibility –aging

10. Silk Fibroin Webs of insects and spiders Antiparallel  -sheets –Rich in Ala and Gly –Close packing of  -sheets –H-bonding between all backbone N-H and C=O Extended but flexible

12. Globular proteins Variety of structures/functions –Enzymes, transport proteins, motor, regulatory, immunoglobulin Folding is compact –H philic  outside –H phobic  inside Human serum albumin Alcohol dehydrogenase N-acetylglucosamine acyltransferase

13. Globular proteins are very compact 3° structure

14. How is the 3D structure determined? X-ray crystallography –Form ‘crystals’ of the protein Regularly repeating lattice X-ray beam is diffracted by the lattice Just like a microscope –Much shorter wavelength (higher energy) light –Computer acts as a ‘lens’ Size of protein is theoretically unlimited

16. How is the 3D structure determined? X-ray crystallography –Get a ‘snapshot’ of the protein in a solid-ish phase –Need highly ordered crystals –Proteins come in close contact: may influence the structure

17. How is the 3D structure determined? NMR –Nuclear spins of 1 H, 13 C, 15 N, etc. Detect via energetic response to a magnetic field Response depends on chemical environment –Distance between all pairs of atoms within the molecule –Software (with plenty of help from the user) determines structures that satisfy these distances

19. How is the 3D structure determined? NMR –Only fairly small (<25kDa) proteins –Need highly concentrated sample Lots of protein Very soluble NMR and crystallography are complementary techniques