anomalous protein migration SDS-PAGE

Michele Klingbeil mklingb at uoft02.utoledo.edu
Wed Feb 1 15:36:09 EST 1995


In article <3g8d31$t46 at israel-info.datasrv.co.il> Lenny Garfinkel,
lenny at zeus.datasrv.co.il writes:
> When I add a pure protein to the sup and incubate for anywhere from 0.5
hr to   >overnight, I see an apparent INCREASE in molecular weight on
SDS-PAGE in the >presence of mercaptoethanol (yup, it runs slower).  What
sort of protein
>modifications could cause this?  Could clipping of a few residues result
>in less SDS binding?

I believe that hydrophobic regions of a polypeptide bind more SDS than 
hydrophilic regions.  So, if your proteolysis cleaved a number of
hydrophobic 
residues than you might see a slower mobility in SDS-PAGE.  If  I remember
correctly you should be able to find some additional information and
possibly
some references in a book titled "Protein Structure."  This book is part
of the
practical approach series and this volume  was edited by T.E. Creighton.  

I would also like to post a question of my own.  I work on the multienzyme
complex, the pyruvate dehydrogenase complex.  I have been studying the 
subunit composition and altered regulatory properties of this complex
from an
ANAEROBIC source.  I also see a change in migration of one of the
subunits.
However, this change in mobility is associated with the phosphorylation
of the
E1 aplha subunit.   This subunit incorporates about 2 mol of 32P/ mol of
E1 aplha,
and I was surprised to see a change in mobility of 2 kDa.  The
unphosphorylated 
subunit migrates at 41 kDa and the phosphorylated form migrates at 43 kDa
using 
SDS-PAGE.  I also know that this subunit has a chnage in pI upon
phosphorylation 
as well ( from 6.8 for the unphosphoryalted subunit to about 6.05 for the
phosphorylated form). Can anyone offer a good explanation for this change
in
mobility?

_____________________________________________
Michele M. Klingbeil						mklingb at uoft02.utoledo.edu	
Dept. of Biology								     	 office:  419 - 537 - 4586		               
      
University of Toledo            fax:      419 - 537 - 7737
Toledo, OH 43606
_____________________________________________



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