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1x1t
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(New page: 200px<br /><applet load="1x1t" size="450" color="white" frame="true" align="right" spinBox="true" caption="1x1t, resolution 1.52Å" /> '''Crystal Structure of...)
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Revision as of 23:09, 24 November 2007
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Crystal Structure of D-3-Hydroxybutyrate Dehydrogenase from Pseudomonas fragi Complexed with NAD+
Overview
The gene coding for d-3-hydroxybutyrate dehydrogenase (HBDH) was cloned, from Pseudomonas fragi. The nucleotide sequence contained a 780 bp open, reading frame encoding a 260 amino acid residue protein. The recombinant, enzyme was efficiently expressed in Escherichia coli cells harboring, pHBDH11 and was purified to homogeneity as judged by SDS-PAGE. The enzyme, showed a strict stereospecificity to the D-enantiomer (3R-configuration), of 3-hydroxybutyrate as a substrate. Crystals of the ligand-free HBDH and, of the enzyme-NAD+ complex were obtained using the hanging-drop, vapor-diffusion method. The crystal structure of the HBDH was solved by, the multiwavelength anomalous diffraction method using the, SeMet-substituted enzyme and was refined to 2.0 A resolution. The overall, structure of P.fragi HBDH, including the catalytic tetrad of Asn114, Ser142, Tyr155, and Lys159, shows obvious relationships with other members, of the short-chain dehydrogenase/reductase (SDR) family. A cacodylate, anion was observed in both the ligand-free enzyme and the enzyme-NAD+, complex, and was located near the catalytic tetrad. It was shown that the, cacodylate inhibited the NAD+-dependent D-3-hydroxybutyrate, dehydrogenation competitively, with a Ki value of 5.6 mM. From the, interactions between cacodylate and the enzyme, it is predicted that, substrate specificity is achieved through the recognition of the 3-methyl, and carboxyl groups of the substrate.
About this Structure
1X1T is a Single protein structure of sequence from Pseudomonas fragi with MG, CAC and NAD as ligands. Active as 3-hydroxybutyrate dehydrogenase, with EC number 1.1.1.30 Full crystallographic information is available from OCA.
Reference
D-3-hydroxybutyrate dehydrogenase from Pseudomonas fragi: molecular cloning of the enzyme gene and crystal structure of the enzyme., Ito K, Nakajima Y, Ichihara E, Ogawa K, Katayama N, Nakashima K, Yoshimoto T, J Mol Biol. 2006 Jan 27;355(4):722-33. Epub 2005 Nov 14. PMID:16325199
Page seeded by OCA on Sun Nov 25 01:17:10 2007
Categories: 3-hydroxybutyrate dehydrogenase | Pseudomonas fragi | Single protein | Ichihara, E. | Ito, K. | Nakajima, Y. | Ogawa, K. | Yoshimoto, T. | CAC | MG | NAD | Beta-hydroxybutyrate | Ketone body | Nad | Nadh | Sdr | Short chain dehydrogenase
