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PDBsum entry 1db3
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Structural and kinetic analysis of escherichia coli gdp-Mannose 4,6 dehydratase provides insights into the enzyme'S catalytic mechanism and regulation by gdp-Fucose.
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Authors
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J.R.Somoza,
S.Menon,
H.Schmidt,
D.Joseph-Mccarthy,
A.Dessen,
M.L.Stahl,
W.S.Somers,
F.X.Sullivan.
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Ref.
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Structure, 2000,
8,
123-135.
[DOI no: ]
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PubMed id
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Abstract
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Background: GDP-mannose 4,6 dehydratase (GMD) catalyzes the conversion of
GDP-(D)-mannose to GDP-4-keto, 6-deoxy-(D)-mannose. This is the first and
regulatory step in the de novo biosynthesis of GDP-(L)-fucose. Fucose forms part
of a number of glycoconjugates, including the ABO blood groups and the selectin
ligand sialyl Lewis X. Defects in GDP-fucose metabolism have been linked to
leukocyte adhesion deficiency type II (LADII). Results: The structure of the
GDP-mannose 4,6 dehydratase apo enzyme has been determined and refined using
data to 2.3 A resolution. GMD is a homodimeric protein with each monomer
composed of two domains. The larger N-terminal domain binds the NADP(H) cofactor
in a classical Rossmann fold and the C-terminal domain harbors the
sugar-nucleotide binding site. We have determined the GMD dissociation constants
for NADP, NADPH and GDP-mannose. Each GMD monomer binds one cofactor and one
substrate molecule, suggesting that both subunits are catalytically competent.
GDP-fucose acts as a competitive inhibitor, suggesting that it binds to the same
site as GDP-mannose, providing a mechanism for the feedback inhibition of fucose
biosynthesis. Conclusions: The X-ray structure of GMD reveals that it is a
member of the short-chain dehydrogenase/reductase (SDR) family of proteins. We
have modeled the binding of NADP and GDP-mannose to the enzyme and mutated four
of the active-site residues to determine their function. The combined modeling
and mutagenesis data suggests that at position 133 threonine substitutes serine
as part of the serine-tyrosine-lysine catalytic triad common to the SDR family
and Glu 135 functions as an active-site base.
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Figure 1.
Figure 1. GDP-fucose biosynthesis. NADP bound to GMD is
reduced and then oxidized during the course of the reaction. GFS
catalyzes two distinct reactions: the epimerization of the
GDP-4-keto, 6-deoxymannose at C3 and C5 followed by the
subsequent reduction at C4 to yield GDP-fucose.
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The above figure is
reprinted
by permission from Cell Press:
Structure
(2000,
8,
123-135)
copyright 2000.
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