Re: AAA problem

Sandra Smith (sandie@mail.utexas.edu)
Fri, 6 Nov 1998 15:48:51 -0500 (EST)

>In the late 80s Speicher reported (Protein Society Meeting poster?) the use
>of DMSO for quantitative determination of cysteine in proteins/peptides.
>The method is based on conversion of cysteine to cysteic acid. This
>conversion happens during the gas phase hydrolysis with the use of HCl
>which contains 4% DMSO. The method works great for cysteines, but affects
>many other AA (Met being one of the most affected - oxidation). For that
>reason I suspect that for the "regular" amino acid analysis any DMSO
>present in the sample has to be completely removed prior to hydrolysis,
>This may be a nontrivial task requiring vacumm drying and redrying of the
>sample until all the DMSO is removed.
>
>Jacek Mozdzanowski
>Bioanalytical Sciences
>SmithKline Beecham

Dear Roger,

With DMSO hydrolsis(liquid phase), the AA's negatively affected are tyr,
his, met, and to a lesser extent ser, thr, and proline. However, if you
know the composition of the peptide and are only interested in
concentration I think you could establish that using the ratios of the
unaffected AA's. I used to use asp. ala and leu when analysing for cysteic
acid. If you can establish the # of pmoles for one residue (use an
average#) then just multiply by the # of residues for total pmoles of each
AA,convert to grams and add back H2O. I'm sure there's some level of error
but I would guess that it would be well under a microgram for the amt
loaded on the average AAanalyser.
The reference for the use of dmso in hydrolysing proteins that I'm aware of is:

Anal Biochem 32, 185-189 (1969)

Sandie Smith
Research Associate
Protein Microanalysis Facility,
Institute for Cellular and Molecular Biology
University of Texas, Austin, Texas 78712
Email: sandie@mail.utexas.edu
Phone: (512) 471-3741, Fax: 512) 471-2149