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Fidelity and Promiscuity of a Mycobacterial Glycosyltransferase
journal contribution
posted on 2016-06-14, 00:00 authored by Kenzo Yamatsugu, Rebecca A. Splain, Laura L. KiesslingMembers
of the genus Mycobacterium cause devastating
human diseases, including tuberculosis. Mycobacterium tuberculosis can resist some antibiotics because of its durable and impermeable
cell envelope. This barrier is assembled from saccharide building
blocks not found in mammals, including galactofuranose (Galf). Within the cell envelope, Galf residues
are linked together to afford an essential polysaccharide, termed
the galactan. The formation of this polymer is catalyzed by the glycosyltransferase
GlfT2, a processive carbohydrate polymerase, which generates a sequence-specific
polysaccharide with alternating regioisomeric β(1–5)
and β(1–6) Galf linkages. GlfT2 exhibits
high fidelity in linkage formation, as it will terminate polymerization
rather than deviate from its linkage pattern. These findings suggest
that GlfT2 would prefer an acceptor with a canonical alternating β(1–5)
and β(1–6) Galf sequence. To test this
hypothesis, we devised a synthetic route to assemble oligosaccharides
with natural and non-natural sequences. GlfT2 could elongate each
of these acceptors, even those with non-natural linkage patterns.
These data indicate that the glycosyltransferase is surprisingly
promiscuous in its substrate preferences. However, GlfT2 did favor
some substrates: it preferentially acted on those in which the lipid-bearing
Galf residue was connected to the sequence by a β(1–6)
glycosidic linkage. The finding that the relative positioning of the
lipid and the non-reducing end of the acceptor influences substrate
selectivity is consistent with a role for the lipid in acceptor binding.
The data also suggest that the fidelity of GlfT2 for generating an
alternating β(1–5) and β(1–6) pattern of
Galf residues arises not from preferential substrate
binding but during processive elongation. These observations suggest
that inhibiting the action of GlfT2 will afford changes in cell wall
structure.