Metabolic glycan engineering (MGE) coupled with nitroxide spin-labeling (SL) was utilized to interrogate the heterogeneous environment of cell surface glycans in select cancer and normal cells. This approach exploited the...
A novel method for spin labelling of sialoglycans on the cell surface is described. C9-Azido sialic acid was linked to glycans on live cells via CSTII-catalysed α2,3-sialylation utilizing azido-sialic acid...
Glycosylphosphatidylinositol (GPI) anchorage of cell
surface proteins
to the membrane is biologically important and ubiquitous in eukaryotes.
However, GPIs do not contain long enough lipids to span the entire
membrane bilayer. To transduce binding signals, GPIs must interact
with other membrane components, but such interactions are difficult
to define. Here, a new method was developed to explore GPI-interacting
membrane proteins in live cell with a bifunctional analogue of the
glucosaminylphosphatidylinositol motif conserved in all GPIs as a
probe. This probe contained a diazirine functionality in the lipid
and an alkynyl group on the glucosamine residue to respectively facilitate
the cross-linkage of GPI-binding membrane proteins with the probe
upon photoactivation and then the installation of biotin to the cross-linked
proteins via a click reaction for affinity-based protein isolation
and analysis. Profiling the proteins pulled down from the Hela cells
revealed 94 unique and 18 overrepresented proteins compared to the
control, and most of them are membrane proteins and many are GPI-related.
The results have proved not only the concept of using the new bifunctional
GPI probe to investigate GPI-binding membrane proteins but also the
important role of inositol in the biological functions of GPI anchors
and GPI-anchored proteins.
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