1. Couple of lab things 1.In calculating rate (  mol/min/mg) from absorbance/time slope, take the absolute value (rates always (usually?) positive) 2.For write-up: in your table of rates, please include the raw data (  abs/time) as well as the calculated rate

2. + + + + + - - - 0 0 0 0 +2 +1 0 +1.5 +0.5 -0.5 pI between pK R and pK 2

3. Free amino acid vs. polymers Pentapeptide (five  carbons!) What’s the charge at pH=7?

4. Different polypeptides have different charge characteristics 1.Amino terminus (if ‘free’) 2.Carboxyl terminus 3.Charged side chains Calculated pI ~7.0 http://www.embl-heidelberg.de/cgi/pi-wrapper.pl

5. Free amino acids vs. polymerized –Side chains may have different pK a s pKa affected by charges on amino/carboxyl groups pKa may be affected by interactions with other side chains in the larger molecule

6. Nucleophilic B: strengthens the acidic character of the adjacent serine

7. Look at protein structure at a low resolution Primary (1°) structure –Sequence of amino acids Total number: anywhere from two to tens of thousands –Few (2 to tens) amino acids: oligopeptides »Typically hormones, etc. –Hundreds or more »“Proteins” »Enzymes, structural proteins, etc.

8. Look at protein structure at a low resolution Non-amino acid chemical groups can enhance protein function –“Prosthetic groups” “Enzyme co-factor” –Associated or covalently-bound –eg. Metals Iron, Calcium, Zinc, Magnesium, etc. Structural components Good nucleophiles: enzymatic ‘activation’ of water, for example Redox chemistry: accept/donate electrons

9. Look at protein structure at a low resolution Prosthetic groups –Lipids (lipoproteins) or sugars (glycoproteins) Enhance protein stability Alter interactions with other biological molecules http://www.liv.ac.uk/physiology/ncs/conform.html

10. Look at protein structure at a low resolution Primary (1°) structure Secondary (2°) structure –Arrangement of local stretches of amino acids –Stabilized predominantly by hydrogen bonds between backbone N-H and O=C

11. Common elements of 2° structure  helix  strand

12. Look at protein structure at a low resolution Primary (1°) structure Secondary (2°) structure Tertiary (3°) structure –Three-dimensional fold of a polypeptide –How do 2° structure elements interact? Non-covalent interactions –Hydrophobic interactions –H-bonds –van der Waals Covalent bonds: disulfide

13.  -helices interact to give the overall 3D fold of a polypeptide

14. Look at protein structure at a low resolution Primary (1°) structure Secondary (2°) structure Tertiary (3°) structure Quaternary (4°) structure –Interaction between subunits of a multi- subunit protein Non-covalent interactions Disulfide (covalent) bonds

15. PROTEIN SUBUNITS (INDIVIDUAL POLYPEPTIDES)

16. Different proteins have different chemical characteristics Caused by 1°, 2°, 3°, 4° structures Define their biological roles Can be exploited to separate proteins from each other –Purify a single protein of interest

17. Different proteins have different chemical characteristics 1.Charge Isoelectric point At pH > pI, net charge ? 0 At pH < pI, net charge ? 0 2.Size Sum of masses of all amino acids (minus 18) Effective size can be influenced by 3°, 4° structure

18. Effective sizes can be influenced by protein shape Globular Filamentous

19. Different proteins have different chemical characteristics 3.Ligand-binding/Affinity for other molecules 4.Exposed hydrophobic patches

20. Exploitation of chemical characteristics Purification –Chromatography Separation/analysis –Electrophoresis