[RASMB] membrane protein

[Jo Butler]

Dear Songpon,

Neal Robinson gives a good description of one approach to handling 
detergent which has been used to solubilise membrane proteins, and it is a 
widely used one.
However, I would like to put in a word for an alternative which does not 
depend upon characterising the detergent, namely measuring the density 
increment of the protein as it occurs at constant chemical potential of all 
diffusible components (i.e. effectively at dialysis equilibrium or as it 
comes off a gel permeation column in the relevant buffer).  This technique 
is based on the fundamental work of Casassa & Eisenberg, H. (1964; 
Thermodynamic analysis of multicomponent solutions, Adv Prot Chem 19, 
287-395) and, if the protein concentration is measured alone (e.g. by amino 
acid analysis) will give the molecular mass of the protein component of any 
micelle - which is often what one actually wants - without involving any 
approximation.
Measuring density increments is not difficult, with the Paar Density meter 
one needs <1ml of solution (and solvent), preferably at a concentration of 
a few mg/ml - but the material is recovered and is available for the 
sedimentation runs.
I have applied this technique to a number of proteins, describing it at a 
Meeting of the UK AUC Users Group (Butler; 1998; Use of density increment 
in assessing protein aggregation under unusual conditions, Biochem Soc 
Trans 26, 749-753).  It has the great advantage that one is actually 
measuring the relevant parameters to get the answer which you actually want 
in most cases.

Jo Butler

--On Tuesday, February 12, 2002 6:24 pm -0800 Songpon Deechongkit 
<songpon at scripps.edu> wrote:

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> Hi all:
>
> I have a few question regarding running AUC on membrane protein.  I would
> appreciate if you can spare your time to help me.
>
> 1) Is it possible to get good/reliable result from membrand protein?
> 2) What are concerns or rules that I should think about when I set up the
> run?
> 3) How much detergent can there be present in the sample, if at all?
>
> Once again, I really appreciate your time.
>
> Sincerely,
> Songpon Deechongkit
>
> The Scripps Research Institute
> La Jolla, California
>
>
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P.J.G. Butler,
MRC Laboratory of Molecular Biology,
Hills Road, Cambridge, CB2 2QH, UK.
Tel. +44 (0)1223 402296