From: Jo Butler <pjgb@mrc-lmb.cam.ac.uk>
  To  : Bogos Agianian <b.agianian@chem.uu.nl>
  Date: Tue, 26 Jan 1999 17:44:18 +0000

Re: AUC and detergents

Dear Bogos,

A good way of handling the problem of determining the protein aggregation
state, at least for sedimentation equilibrium, is to calculate the apparent
partial specific volume of the protein/detergent micelle from the measured
density increment.  If this latter is determined using the PROTEIN
concentration (i.e. without considering the bound detergent), the classic
work of Casassa and Eisenberg shows that you will get a molecular mass
corresponding to the protein component alone.  This result is absolute and
not an approximation, provided the solution and solvent were at constant
chemical potential of all diffusible components, i.e. at dialysis
equilibrium, before the density measurements and analytical centrifugation.
I discussed this use (among others) of measuring density increments at the
1998 UK Analytical Users Group Meeting in Nottingham, and the paper was
recently published (Butler, P. J. G. (1998). Use of density increment in
assessing protein aggregation under unusual conditions. Biochem. Soc.
Trans. 26, 749-753).

I hope that this information is helpful,

Jo Butler

--On Tue, Jan 26, 1999 2:30 pm +0100 Bogos Agianian <b.agianian@chem.uu.nl>
wrote:

> Dear RASMB users,
> 
> We wish to examine several different protein samples using AUC (both
> sedimentation velocity and equilibrium), all of which contain
> detergent(s) at concentrations below or above their CMC depending on
> the particular protein of study.  I would appreciate your expert
> opinion on the effect of the presence of detergent on AUC data
> interpretation and analysis. More specifically, I would like to ask:
> 
> 1. How much do detergents affect determination of the s and MW values
> and which are the corrections one should apply to get the 'true'
> values out
> 
> 2. How should one handle data (both from velocity and equilibrium
> runs) from samples that contain detergent micelles?
> 
> Thank you in advance for your response
> 
> Bogos Agianian
> 
> 
> 
> 
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Dr P.J.G. Butler,
MRC Laboratory of Molecular Biology,
Hills Road,
Cambridge, CB2 2QH,
UK.
Tel. 01223 248011 (or 01223 402296 DDI)
FAX. 01223 213556