From: Diane Wakeham <wakeham@itsa.ucsf.edu>
  To  : rasmb@alpha.bbri.org
  Date: Tue, 12 Sep 2000 18:40:00 -0700 (PDT)

Re: MW Determination for Rod- Shaped Protein(Axial Ratio?)

RASMBers,

Many thanks for your suggestions.

My protein monomer is 52 kD and pI is 6.14.  I calculated vbar using
Ultrascan as 0.732 from the sequence, so that's not a significant
contribution to the error.  I do believe that my rough estimate of axial
ratio is way too low (perhaps, since it's essentially cylindrical, I
should take length/radius and get an axial ratio of 8.5?  I plan to try to
use hydropro or ellipsprime and ellips1 - or other programs you suggest?-
to give me a better estimate starting from known .pdb of a twenty short
helix fragment that includes most of the protein, since it's fairly
rigid).

Data gathered at a range of pH values suggests the charge could be the
largest component of nonideality, because MW quite close to the actual
value was measured at pH 6.2, with a high dependence on the buffer
solution chosen.   I'm now doing an experiment at different concentrations
(I started at 0.1 mg/mL, likely below Kd for dimerization) and will follow
your suggestions using Nonlin for fitting the data.  I appreciate all your
advice!

Diane Wakeham