1. RE digests & RE maps OD and uses Last class How to read a paper Intro to Paper 1

2. Purifying proteins Purification – based on what properties?

3. Purifying proteins - Solubility [Salt] Fraction protein in precipitate + + + + - - - - + + + + - - - - + + + + - - - - + + + + - - - -

4. Ammonium sulfate precipitation

5. Getting rid of salt: Dialysis

6. Ammonium sulfate precipitation 0-40% 40-60% 60-80% Bradford assay & enzyme assay (why)? - Every fraction Lab Dialysis

7. First evidence of PA-GFP…

8. Increase in fluorescence of PA-GFP…

9. pH affects

10. Photostability

11. OK – what about in cells?

12. Can we study dynamics?

13. Lets do a real experiment – live trafficking

14. Following live trafficking of proteins

15. Lysosomal trafficking needs microtubules