RE: MS of deglycosylated proteins
Dirk S. Krapf (dkrapf@gilson.com)
Tue, 23 Mar 1999 19:57:23 -0600
>Date: Tue, 23 Mar 1999 10:25:26 -0800
>From: Viswanatham Katta <vkatta@amgen.com>
>
>Sub: Reply to MS of deglycosylated proteins
>
>Hi Dougie
>
>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
>
>----------------------------------------------------------------------------
>In reply to Dougie Lamont's question:
>
>
>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
>
>
>