SO YOU WANT TO USE THE BIOPHYSICS PLATFORM?

You might want to first chat with Mike Osborne or Katherine Borden

1. Nearly all work is done in-vitro

2. You need pure samples (> 95% pure from SDS-page gel)

3. You will likely need quite a lot of it (we recommend, and can help, expressing in E. coli).

4. For Fluorescence and Circular Dichroism you need ~ 1-10mircoM in 200 microL of protein

5. For AUC, which is an OD-based technique, you need ~OD280 = 0.4 and 400 microL

6. For NMR you will need > 50 microM protein in ~ 300 microL

i. Note for structure determination, there is a Mwt limit of ~ 30-35 kDa

ii. For structure you will need to express with stable 15N/13C isotopes (we have protocols for this)

Some basic uses of the techniques are:

Fluorescence: protein ligand interactions, folding/unfolding, Kd, stoichiometry

CD: monitor protein structure, secondary structure content, effect of mutants, interactions

NMR: structure determination, protein motions, mapping of binding sites