Re: Question
Gregory Grant (ggrant@pharmdec.wustl.edu)
Fri, 5 Feb 1999 09:28:34 -0600
>Hi,
>
>My name is Eric Tam and I'm a biochemistry PhD student here at the
>University of British Columbia in Vancouver, Canada. I was recommended this
>address by the Nucleic Acids and Protein Services (NAPS) of UBC. I have a
>question regarding biotin modification and trypsin digestion.
>
>If a primary amine is modified by a biotinylation reagent (say from Pierce),
>is trypsin able to cleave at this modified site? The reason I'm asking this
>is that I'm proposing to biotinylate a protein to identify surface exposed
>Lys residues. I then want to be able to digest the protein and sequence
>only the biotinylated peptides.
>
>Your response is greatly appreciated,
>
>Eric Tam
Eric,
If I understand correctly what you are doing, the modified lysine
side chains will no longer be recognized by trypsin. A free primary amine
is necessary for this. So you will have to use another protease for the
digestion. Such as chymotrypsin, Glu-C, Arg-C, Asp-N, etc. I don't see this
as a problem and it should work fine. The biotinylated peptides can be
iosolated by affinity chromatography.
Gregory A. Grant
ggrant@pharmdec.wustl.edu