[RASMB] Equilibrium runs with small peptides, high offset values

[Joachim Behlke]

Von:            	Erin Matthews <erin.matthews at yale.edu>
Organisation:   	Yale University
An:             	rasmb at rasmb-email.bbri.org
Betreff:        	[RASMB] Equilibrium runs with small peptides, high offset values
Datum:   	Mon, 09 Dec 2002 19:34:43 -0500

Erin,

As Peter Schuck has recommended small peptides can be 
analyzed well in sedimentation velocity experiments. Some years 
ago we have studied succesfully angiotensin I and II with a 
molecular mass of about 1.2 or 1.0 kDa using the whole boundary 
sedimentation and fitting by an approximate solution of the Lamm 
equation (see J. Behlke and O. Ristau: Biophys.Chem.70, 133-146 
1998). The program Lamm which we have improved recently is 
available on the RASMB site. 
Best wishes.
Joachim Behlke
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> 
> Thanks everyone for suggestions on my last question about data with
> negative slopes!  I have yet another question on a different system.
> 
> I did an equilbrium (absorbance optics) run on a water   
> soluble  peptide of 1290 Daltons.  I had to spin at 60K rpm 
> to get any reasonble data (data at lower speeds showed virtually no 
> curvature).  My problem now is this: using Nonlin,  the best fit to the
> data gives a lower than expected sigma (.35 instead of .46 with both the 
> offset and the second virial coefficent set to 0).  There doesn't appear 
> to be a higher order association.  When I fix sigma to the 'known'
> value, the fit is poor, but when I then let the offset float I get a
> good fit, but the offset is .1332.  Very high!  I obviously can't do an
> overspeeding experiment to get an experimental measure of the depleted
> meniscus absorbance.  As well, I imagine that the calculated partical
> specific volume (from Durchschlag and Zipper, 1994) could be inaccurate.
> Is there something more I can be doing to get an accurate fit?  Which
> would be considered a more physically reasonble fit to the data--a fit
> using a lower than expected sigma value or a high offset? 
> 
> Thanks,
> 
> Erin Matthews
> Graduate Student, Engelman Lab, Yale University
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