IUBio

SDS-PAGE migration of lipoproteins

John A. Newitt newitt at removeme.nih.gov
Mon Mar 15 14:21:11 EST 1999


In article <v03102802b309bd4aa31a@[128.135.42.13]>,
cbookste at MEDICINE.BSD.UCHICAGO.EDU (Cres Bookstein) wrote:

> Funny you should ask.  We have been discussing the unusual mobility on
> SDS-PAGE of a mammalian membrane protein with heating at 65oC vs boiling.
> This protein is not covalently bound to any lipid moiety.  It has 10-12
> membrane spans and a large soluble domain. With boiling, the antigenic band
> runs at approximately 200 kDaltons.  At 65oC or less, it runs at the
> molecular weight predicted by amino acid sequence.   Another protein that
> behaves like this is one of the glucose transporters (Glut5?), also with
> 10-12 membrane spans and a large soluble domain and no lipid moieties.
> Our explanation is purely guess...that the boiling causes aggregation; the
> lower temperature allows unfolding without aggregation. Similar experiences
> of others will be watched with interest.
> 

This is actually not an uncommon observation with membrane proteins. For
this reason, and for the fact that some Asp-Pro bonds appear to be labile
when boiling in SDS-PAGE sample buffer, I always have prepared my samples
for SDS-PAGE by heating at 65 degrees C for 30 minutes.

Regards,

John A. Newitt, Ph.D.           |   <newitt at removeme.nih.gov>
National Institutes of Health   |   FAX: 301-402-0387
Bethesda, Maryland  USA         |   

Please delete "removeme." from my address to reply by email.



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