Dear Linda,
Further to the wisdom below, in many cases when I have collected data that
is consistent with sequencing through an N-linked glycosylation site, there
is evidence for the predicted/expected Asn as a very weak, but real Asp, is
observed.
Regards,
alex
Alexander Bell
Anat and Cell Biol
McGill University
Montreal, Quebec, Canada
----- Original Message -----
From: "David C. Chiara" <david_chiara@hms.harvard.edu>
To: "Recipients of ABRF List" <abrf@aecom.yu.edu>
Cc: "Recipients of ABRF List" <abrf@aecom.yu.edu>
Sent: Tuesday, November 21, 2000 8:49 AM
Subject: Re: Effect of glycosylation on sequencing
> Linda,
>
> I can only comment on N-linked glycosylation. The glycosylation site does
> not hinder sequencing although you will not see the ASN residue in that
> cycle. Pre-treatment with N-glycanase or PGNase F will remove the
> carbohydrate and modify the ASN to an ASP residue. If the glycosylation
> site is high mannose (or some hybrids) and is cleaved by endoglycosidase
H,
> the resulting amino acid is N-acetylglucosamine asparagine who's PTH
> derivative elutes between DTT and ASP-PTH. See Paxton et al, (1987)
> Biochemistry 84, 920-924.
>
>
> David C. Chiara, M.D., Ph.D.
> Research Associate
> Department of Neurobiology
> Harvard Medical School
> 220 Longwood Avenue
> Boston, Ma. 02115
> ph: (617) 432-1729
> fax: (617) 734-7557
> Email: David_Chiara@hms.harvard.edu
> www.hms.harvard.edu/bss/neuro/cohen/
>
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