We have successfully used MALDI-TOFMS (Voyager DE-RP, PE Biosystems Inc.) a
number of times to determine the protein backbone mass after
de-N-glycosylation. We were able to answer questions about the correct
processing of the N-terminus or C-terminus of the protein. One advantage of
this approach is that the deglycosylation doesn't need to go to completion.
Even with about 20% of the protein is completely de-N-glycosylated (rest of
the protein deglycosylated to various extents) we can get good MW of the
backbone of the protein. Another advantage is no chromatographic separation
is required.
Viswanatham Katta
Protein structure Dept.
Amgen Inc.
e-mail: vkatta@amgen.com
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hi all,
we have a single quad from sciex (API100) and have been looking at methods
to determine the mass of the protein backbone (40-80KDa) after
deglycosylation. I wondered what the viewpoint was of using electrospray for
this type of work or would MALDI-MS be more appropriate. Also we have tried
a number of methods to deglycosylate and have found that detergents appear
to be necessary for complete removal of glycans. Any views or opinions on
this would be welcome.
I'm off to the ABRF meeting in North Carolina in a few days and would like
to take the opportunity to speak to anybody who has experience in this area.
If you can offer any advice in this area and would like to chat it would be
much appreciated.
yours
Dougie lamont
PPL Therapeutics