From: Arthur Rowe <Arthur.Rowe@nottingham.ac.uk>
  To  : libby@groucho.med.jhmi.edu, RASMB@bbri.org
  Date: Tue, 16 Nov 1999 12:11:46 GMT0BST

Re: s coeff and shape

Dear Libby

The method is simple enough, in principle.  Find the frictional ratio (f/f0) using 
SEDNTERP, using your M value, an s value, and vbar if you have it (taking it as 
0.73 ml/g will be good enough if you don't.  Or, of course, use a picket calculator 
and the relevant equation.

If f/f0 is significantly greater than 1.2, you can be confident that your protein is 
either (i) assymetric, or (ii) swollen, with a substantial internal cavity filled with 
water.  The latter is only really likely with a large, oligomeric structure (e.g. a 
multi-enzyme complex).

As regards that s value.  If you are working at around (say) 1 mg/ml, then do not 
bother about any c-dependence.  Just take the s value at the maximum in your g(s*) 
profile. You are looking for serious assymetry, I gather, so who cares if the s value 
requires correction by the odd percent, or two ?  

I think, incidentally, that you are mis-interpreting your van Holde/Weischet plot - 
I am not aware that it can readily give you a value for ks (the c-dependence 
coefficient).  If you actually want to check  c-dependence out, its best to do multiplex 
3 cells at 3 starting values of c, and find the linear dependence of s on c from the 
g(s*)max values plotted against c (corrected for radial dilution).  ks values of 
around 8 ml/g or upwards are again indicative of either assymetry or swelling.

All best

Arthur Rowe


you wrote:

 We are working on a protein that we believe is asymmetric.  I'm trying to get 
sedimentation velocity data to hopefully support this hypothesis.  I do NOT need 
an exact, definitive shape, all I am interested in is knowing that our proposed 
(asymmetric) model is one way of interpreting the sedimentation coefficient. 

 I know the following: 

 1.  protein is a single species by sed. eq. 2.  s shows concentration dependence (2.5-4 
S) by vanHolde-Weischet analysis 

 My questions:  

 What analysis method(s) is the best for my problem ??  

 If s is concentration dependent, may I take an average s from a g(s) analysis and 
make crude shape predications based on that ?? 

 Thanks in advance for your advice, 

 Libby 

 

 
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