1r66

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Revision as of 20:05, 24 November 2007


1r66, resolution 1.44Å

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Crystal Structure of DesIV (dTDP-glucose 4,6-dehydratase) from Streptomyces venezuelae with NAD and TYD bound

Overview

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.

About this Structure

1R66 is a Single protein structure of sequence from Streptomyces venezuelae with CL, NAD and TYD as ligands. Active as dTDP-glucose 4,6-dehydratase, with EC number 4.2.1.46 Full crystallographic information is available from OCA.

Reference

High resolution X-ray structure of dTDP-glucose 4,6-dehydratase from Streptomyces venezuelae., Allard ST, Cleland WW, Holden HM, J Biol Chem. 2004 Jan 16;279(3):2211-20. Epub 2003 Oct 21. PMID:14570895

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