Synthesis
of Biologically Active <i>N</i>- and <i>O</i>‑Linked
Glycans with Multisialylated Poly‑<i>N</i>‑acetyllactosamine
Extensions Using <i>P. damsela</i> α2‑6 Sialyltransferase
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Abstract
Sialosides
on <i>N</i>- and <i>O</i>-linked glycoproteins
play a fundamental role in many biological processes, and synthetic
glycan probes have proven to be valuable tools for elucidating these
functions. Though sialic acids are typically found α2-3- or
α2-6-linked to a terminal nonreducing end galactose, poly-LacNAc
extended core-3 <i>O</i>-linked glycans isolated from rat
salivary glands and human colonic mucins have been reported to contain
multiple internal Neu5Acα2-6Gal epitopes. Here, we have developed
an efficient approach for the synthesis of a library of <i>N</i>- and <i>O</i>-linked glycans with multisialylated poly-LacNAc
extensions, including naturally occurring multisialylated core-3 <i>O</i>-linked glycans. We have found that a recombinant α2-6
sialyltransferase from <i>Photobacterium damsela</i> (Pd2,6ST)
exhibits unique regioselectivity and is able to sialylate internal
galactose residues in poly-LacNAc extended glycans which was confirmed
by MS/MS analysis. Using a glycan microarray displaying this library,
we found that Neu5Acα2-6Gal specific influenza virus hemagglutinins,
siglecs, and plant lectins are largely unaffected by adjacent internal
sialylation, and in several cases the internal sialic acids are recognized
as ligands. Polyclonal IgY antibodies specific for internal sialoside
epitopes were elicited in inoculated chickens