Hi everyone,
Recently, I obtained a soluble glyco-protein. Unfortunately, after I added
PNGase or Endo Hf to remove the glycans, the deglycosylated protein is
precipitated. Is there any method to avoid this kind of precipitation?
Thanks,
Simon
Hi Simon,
Although they are a source of heterogeneity for
crystallization, glycosylations usually stabilize proteins.
There are a couple of things that may be important to consider before you
deglycosylate your protein.
Do you know how many natural sequons/glycosylation sites your protein has?
My apologies Simon, I should have been more thorough answering your
question.
Yes the protein was shown to be quite homogenously glycosylated using
mass-spectrometry.
The ED maps showed the first two ordered fully occupied NAG units and
residual density for a third sugar unit although it is very
Dear Simon,
this may be an isolated case, but you might want to try to drastically
change the pH of the reaction conditions.
In our hands, a protein with 3 hyperglycosylated sites, expressed in Pichia
pastoris, could be deglycosylated readily with endoH. However, at the
recommended reaction pH
