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PDBsum entry 1r6d
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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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High resolution X-Ray structure of dtdp-Glucose 4,6-Dehydratase from streptomyces venezuelae.
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Authors
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S.T.Allard,
W.W.Cleland,
H.M.Holden.
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Ref.
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J Biol Chem, 2004,
279,
2211-2220.
[DOI no: ]
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PubMed id
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Abstract
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Desosamine is a 3-(dimethylamino)-3,4,6-trideoxyhexose found in some macrolide
antibiotics. In Streptomyces venezuelae, there are seven genes required for the
biosynthesis of this unusual sugar. One of the genes, desIV, codes for a
dTDP-glucose 4,6-dehydratase, which is referred to as DesIV. The reaction
mechanisms for these types of dehydratases are quite complicated with proton
abstraction from the sugar 4'-hydroxyl group and hydride transfer to NAD+,
proton abstraction at C-5, and elimination of the hydroxyl group at C-6 of the
sugar, and finally return of a proton to C-5 and a hydride from NADH to C-6.
Here we describe the cloning, overexpression, and purification, and high
resolution x-ray crystallographic analysis to 1.44 A of wild-type DesIV
complexed with dTDP. Additionally, for this study, a double site-directed mutant
protein (D128N/E129Q) was prepared, crystallized as a complex with NAD+ and the
substrate dTDP-glucose and its structure determined to 1.35 A resolution. In
DesIV, the phenolate group of Tyr(151) and O(gamma) of Thr(127) lie at 2.7 and
2.6 A, respectively from the 4'-hydroxyl group of the dTDP-glucose substrate.
The side chain of Asp(128) is in the correct position to function as a general
acid for proton donation to the 6'-hydroxyl group while the side chain of
Glu(129) is ideally situated to serve as the general base for proton abstraction
at C-5. This investigation provides further detailed information for
understanding the exquisite chemistry that occurs in these remarkable enzymes.
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Figure 2.
FIG. 2. Ribbon representation of DesIV. Shown in a is a
ribbon representation of one subunit of DesIV with the bound
ligands, NAD^+ and dTDP, shown in a ball-and-stick
representation. The dimeric form of the enzyme is presented in b.
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Figure 3.
FIG. 3. Close up view of the DesIV active site. A close up
view of the active site, within  3.2 Å of the NAD^+
and dTDP ligands, is shown in a. For the sake of clarity, Ser87
and His88 were omitted from the figure. The side chain hydroxyl
group of Ser87 interacts with a phosphoryl oxygen of the NAD^+
while N  2 of His88 forms a
hydrogen bond with a phosphoryl oxygen of the dTDP. A schematic
of the hydrogen-bonding pattern around dTDP is displayed in b.
Possible hydrogen-bonding interactions within 3.2
Å are indicated by dashed lines.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2004,
279,
2211-2220)
copyright 2004.
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