exoglycosidase activity on native glycoproteins

kzhao at BIOVX1.BIOLOGY.UCLA.EDU kzhao at BIOVX1.BIOLOGY.UCLA.EDU
Thu Jun 19 12:37:26 EST 1997


For David Hoover's question regarding PNGase F deglycosylation:

There is a problem to get native deglycosylated protein using PNGase F, as the
enzyme requires denaturation of glycoproteins to be deglycosylated.  This is 
understandable if you think the position of PNGase F action, i.e., at the base
between sugar chain and the peptide.  Whereas endo-H can work perfectly on n
ative glycoproteins, part of the reason is that it works on GlcNAc-GlcNAc linkage a little away from the base.  Unfortunately, endo-H only works on high mannoseor hybrid structures.

If you are dealing with biantennary complex, I would suggest to use glycopeptidase A (from Seikagaku, very expensive), which is more efficient than PNGase F.

What are the reasons that kept you from using other systems?  I know Genzyme is
using your approach to make their therapeutic recombinant beta-glucosidase for
Gaucher disease.  I'd like to know details of your failed approaches to figure
out additional ways to overcome the problem.

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* Ke-Wei Zhao, Ph.D.                     Phone: (310)825-8722       
* Department of Biological Chemistry     Fax: (310)206-5272
* UCLA School of Medicine, CHS 33-257    e-mail: kzhao at biovx1.biology.ucla.edu
* Los Angeles, CA 90095-1737
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