Deglycosylating (?????) IgG

[Iain Wilson]

ANDREI POPOV <ANDREI.POPOV at bbsrc.ac.uk> wrote:


> <Tunicamycin will not deglycosylate - it merely prevents formation of
> <dolichol-linked oligosaccharides - and so is a biosynthetic inhibitor.
> 
> Thanks for the info
> 
> <The partially-cryptic site of IgG oligosaccharides would create
> <difficulties in using enzymatic deglycosylation, whilst using non-
> <denatured IgG.
> 
> Sorry, but I could not understand this...
> So far I have not heard any complaints about the usage
> of the drug
> 

I thought as a general point I should add something on the difficult
problem that was being posed - hence talk of the cryptic site.

Perhaps to clarify I should say that the proposed crystal structure
of IgG (Sutton and Philips in Nature some years ago) would indicate
that the Fc oligosaccharides are within the Fc cleft. 

Also, I presumed that the original post referred to using normal
IgG - i.e. it comes from a human or other animal rather than from
a cell line. Thus tunicamycin could not be used - the only way to
produce non-glycosylated IgG would be to take purified IgG and find
some way of enzymatically or chemically deglycosylating it. And if 
functional studies were to be performed then the IgG should not be 
denatured - yet for access by deglycosylating enzymes one may need 
the IgG to be denatured due to the restricted site of the 
oligosaccharides: you see the problem? The access problem is 
irrelevant to the question of how effective tunicamycin is.
It would only be relevant to use tunicamycin if you had cell lines
expressing IgG and using tunicamycin as a biosynthetic inhibitor -
the only problem is to what extent normal cell function would be 
affected by a lack of N-linked glycosylation.

Perhaps we confused eachother with the term 'deglycosylated' - to me
it means stuff you remove the glycosylation from - rather than stuff
that has never been glycosylated.

Iain
wilson at edv1.boku.ac.at