unusual carbohydrate structure?

Peter Kulakosky pck3 at CORNELL.EDU
Thu Nov 6 17:27:36 EST 1997


In our  analysis of oligosaccharides from an insect cell viral protein  w=
e
have come across what appears to be an unusual structure.

In an electrophorretic profile, relative to a glucose ladder standard, it
has a size of about 7 GU (glucose units).  When we digest with jack bean
hexosaminidase it shifts downward 1.4 GU to 5.6 GU.  Removal of 1 GlcNAc
usually results in a decrease of 0.75 GU in this system.  So it appears
that 2 GlcNAc=92s were removed.  However, at 5.6 GU this is still larger =
than
the trimannosyl core (4.0 ~ 4.2 GU) or fucosylated trimannosyl core (4.5 =
~
4.7 GU).

When this structure is further digested with jack bean mannosidase no
further digestion results.  The structure remains at 5.6 GU.  Fucosidase
digests remove 1 core fucose, but the structure is still larger by about
1.3 GU than the trimannosyl core.  A combined digest with fucosidase and
hexosaminidase only gave the additive effect of the single digests--no
synergy suggesting outer arm fucose.

The structure is not digested by b-galactosidase or sialidase, nor does
longer exposure to hexosaminidase produce further digestion.  At 5.6 GU
there is still a sugar molecule attached to this structure which prevents
complete digestion by combination of sialidase, b-galactosidase,
hexosaminidase, and a-mannosidase.  We are using Oxford GlycoSystem's
sequencing enzymes.

We have also done monosaccharide analysis on the entire glycoform profile=
,
from which this oligosaccharide was purified, which only revealed the
presence of mannose, GlcNAc, and fucose; there was no indication of
galactose or sialic acid.

Anyone have any suggestions for possible structures?

Thanks




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