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References listed in PDB file
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Key reference
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Title
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Oligosaccharide preferences of beta1,4-Galactosyltransferase-I: crystal structures of met340his mutant of human beta1,4-Galactosyltransferase-I with a pentasaccharide and trisaccharides of the n-Glycan moiety.
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Authors
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V.Ramasamy,
B.Ramakrishnan,
E.Boeggeman,
D.M.Ratner,
P.H.Seeberger,
P.K.Qasba.
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Ref.
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J Mol Biol, 2005,
353,
53-67.
[DOI no: ]
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PubMed id
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Abstract
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beta-1,4-Galactosyltransferase-I (beta4Gal-T1) transfers galactose from
UDP-galactose to N-acetylglucosamine (GlcNAc) residues of the branched N-linked
oligosaccharide chains of glycoproteins. In an N-linked biantennary
oligosaccharide chain, one antenna is attached to the 3-hydroxyl-(1,3-arm), and
the other to the 6-hydroxyl-(1,6-arm) group of mannose, which is beta-1,4-linked
to an N-linked chitobiose, attached to the aspargine residue of a protein. For a
better understanding of the branch specificity of beta4Gal-T1 towards the GlcNAc
residues of N-glycans, we have carried out kinetic and crystallographic studies
with the wild-type human beta4Gal-T1 (h-beta4Gal-T1) and the mutant
Met340His-beta4Gal-T1 (h-M340H-beta4Gal-T1) in complex with a GlcNAc-containing
pentasaccharide and several GlcNAc-containing trisaccharides present in
N-glycans. The oligosaccharides used were: pentasaccharide
GlcNAcbeta1,2-Manalpha1,6 (GlcNAcbeta1,2-Manalpha1,3)Man; the 1,6-arm
trisaccharide, GlcNAcbeta1,2-Manalpha1,6-Manbeta-OR (1,2-1,6-arm); the 1,3-arm
trisaccharides, GlcNAcbeta1,2-Manalpha1,3-Manbeta-OR (1,2-1,3-arm) and
GlcNAcbeta1,4-Manalpha1,3-Manbeta-OR (1,4-1,3-arm); and the trisaccharide
GlcNAcbeta1,4-GlcNAcbeta1,4-GlcNAc (chitotriose). With the wild-type
h-beta4Gal-T1, the K(m) of 1,2-1,6-arm is approximately tenfold lower than for
1,2-1,3-arm and 1,4-1,3-arm, and 22-fold lower than for chitotriose. Crystal
structures of h-M340H-beta4Gal-T1 in complex with the pentasaccharide and
various trisaccharides at 1.9-2.0A resolution showed that beta4Gal-T1 is in a
closed conformation with the oligosaccharide bound to the enzyme, and the
1,2-1,6-arm trisaccharide makes the maximum number of interactions with the
enzyme, which is in concurrence with the lowest K(m) for the trisaccharide.
Present studies suggest that beta4Gal-T1 interacts preferentially with the
1,2-1,6-arm trisaccharide rather than with the 1,2-1,3-arm or 1,4-1,3-arm of a
bi- or tri-antennary oligosaccharide chain of N-glycan.
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Figure 1.
Figure 1. A depiction of complex penta-antennary N-glycan
structure. The pentasaccharide and the trisaccharides of the
N-glycan moiety, used for the kinetic and crystallographic
studies, are highlighted by colored arrows.
GlcNAcb1,2-Mana1,6(GlcNAcb1,2-Mana1,3)-Man pentasaccharide (blue
and green arrows); GlcNAcb1,2-Mana1,6-Manb-OR (1,2-1,6-arm)
(blue arrows); GlcNAcb1,2-Mana1,3-Manb-OR (1,2-1,3-arm) (green
arrows); GlcNAcb1,4-Man a1,3-Manb-OR (1,4-1,3-arm) (purple
arrows).
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Figure 2.
Figure 2. Effects of varying the oligosaccharide acceptor
substrate concentration on the initial rate of galactose
transfer (n) by the h-b4Gal-T1. Chitobiose (GlcNAcb1,4-GlcNAc)
and chitotriose GlcNAcb1,4-GlcNAcb1,4-GlcNAc (0M),
GlcNAcb1,2-Man (  triangle,
open ), GlcNAcb1,2-Mana1,6-Manb-OR (sB),
GlcNAcb1,2-Mana1,3-Manb-OR (  open
), GlcNAcb1,4-Mana1,3-Manb-OR (  triangle,
filled ), pentasaccharide
GlcNAcb1,2-Mana1,6(GlcNAcb1,2-Mana1,3)Man(  ).
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(2005,
353,
53-67)
copyright 2005.
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