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Florescent substrates for glycosyltransferases?

Iain Wilson WILSON at edv1.boku.ac.at
Wed Sep 13 08:08:36 EST 1995


> I have noticed that one can use commercially available AMC-labelled oligosaccharides as substrates 
> for glycosidase assays.
> My question is whether there is any experience around with using similarly labeelled  
> oligosaccharides as substrates for glycosyltransferases. If yes then how far the label should 
> be from the reaction centre and what might be the optimal method for following the reaction, e.g. 
> TLC or HPLC.I am mostly thinking about assaying the activity  of  alpha- galactosyl transferase.
> 
> Thanks,
> Armin Sepp, PhD
> Armin.Sepp at bbsrc.ac.uk

In our lab pyridylamino-oligosaccharides have been used as substrates 
for GlcNAc transferases - e.g. Man5GlcNAc2-PA and Man3GlcNAc2-PA. See 
Altmann et al (1993) Glycobiology 3, 619-625. The oligosaccharides 
were subsequently separated on HPLC (Hypersil ODS) and fluorescence 
was monitored (320/400 nm). If you're thinking of alpha1,3-GalT use 
of an asialo-biantennary N-link with PA label should be possible, 
since GlcNAc-transferases act before alpha1,3GalT. Consider also that 
terminal glycosyltransferases appear tolerant of a range of 
octyloligosaccharides that lack the chitobiose portion of N-linked 
oligosaccharides.

Iain

---------------------------------------------------------------
Iain Wilson                        Institut fuer Chemie           
Tel: 43-1-47654-6065               Universitaet fuer Bodenkultur   
Fax: 43-1-310-5176                 Gregor-Mendel-Strasse 33
E-mail: wilson at edv1.boku.ac.at     A-1180, WIEN, Austria




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