1R6D

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

Summary for 1R6D

• Entry DOI: 10.2210/pdb1r6d/pdb
• Related: 1R66
• Descriptor: TDP-glucose-4,6-dehydratase, NICOTINAMIDE-ADENINE-DINUCLEOTIDE, 2'DEOXY-THYMIDINE-5'-DIPHOSPHO-ALPHA-D-GLUCOSE, ... (4 entities in total)
• Functional Keywords: dehydratase, rossmann fold, short-chain dehydrogenase/reductase, lyase
• Biological source: Streptomyces venezuelae
• Total number of polymer chains: 1
• Total formula weight: 37725.67

Authors

Allard, S.T.M.,Cleland, W.W.,Holden, H.M. (deposition date: 2003-10-14, release date: 2004-01-27, Last modification date: 2023-08-23)

Primary citation

Allard, S.T.M.,Cleland, W.W.,Holden, H.M.
High Resolution X-ray Structure of dTDP-Glucose 4,6-Dehydratase from Streptomyces venezuelae
J.Biol.Chem., 279:2211-2220, 2004

PubMed Abstract: 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.

PubMed: 14570895
DOI: 10.1074/jbc.M310134200

Experimental method

X-RAY DIFFRACTION (1.35 Å)