2rhw

From Proteopedia

Crystal Structure of the S112A mutant of a C-C hydrolase, BphD from Burkholderia xenovorans LB400, in complex with 3,10-Di-Fluoro HOPDA

Overview

The microbial degradation of polychlorinated biphenyls (PCBs) by the, biphenyl catabolic (Bph) pathway is limited in part by the pathway's, fourth enzyme, BphD. BphD catalyzes an unusual carbon-carbon bond, hydrolysis of 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (HOPDA), in, which the substrate is subject to histidine-mediated enol-keto, tautomerization prior to hydrolysis. Chlorinated HOPDAs such as 3-Cl HOPDA, inhibit BphD. Here we report that BphD preferentially hydrolyzed a series, of 3-substituted HOPDAs in the order H > F > Cl > Me, suggesting that, catalysis is affected by steric, not electronic, determinants. Transient, state kinetic studies performed using WT BphD and the hydrolysis-defective, S112A variant indicated that large 3-substituents inhibited, His-265-catalyzed tautomerization by 5 orders of magnitude. Structural, analyses of S112A:3-Cl HOPDA and S112A:3,10-diF HOPDA complexes revealed a, nonproductive binding mode in which the plane defined by the C atoms of, HOPDA's dienoate moiety is nearly orthogonal to that of the proposed keto, tautomer observed in the S112A:HOPDA complex. Moreover, in the 3-Cl HOPDA, complex, the 2-hydroxo group is moved by 3.6 A from its position near the, catalytic His-265 to hydrogen bond with Arg-190 and access of His-265 is, blocked by the 3-Cl substituent. Nonproductive binding may be stabilized, by interactions involving the 3-substituent with non-polar side chains., Solvent molecules have poor access to C6 in the S112A:3-Cl HOPDA, structure, more consistent with hydrolysis occurring via an acyl-enzyme, than a gem-diol intermediate. These results provide insight into, engineering BphD for PCB degradation.

About this Structure

2RHW is a Single protein structure of sequence from Burkholderia xenovorans with , and as ligands. Full crystallographic information is available from OCA.

Reference

The molecular basis for inhibition of BphD, a C-C bond hydrolase involved in polychlorinated biphenyls degradation: Large 3-substituents prevent tautomerization., Bhowmik S, Horsman GP, Bolin JT, Eltis LD, J Biol Chem. 2007 Oct 11;. PMID:17932031

Page seeded by OCA on Wed Jan 23 11:24:32 2008