Abstract

The oxidative degradation of biphenyl and polychlorinated biphenyls (PCBs) is initiated in Pandoraea pnomenusa B-356 by biphenyl dioxygenase (BPDOB356). BPDOB356, a heterohexameric (ab)3 Rieske oxygenase (RO), catalyzes the insertion of dioxygen with stereo- and regioselectivity at the 2,3-carbons of biphenyl, and can transform a broad spectrum of PCB congeners. Here we present the X-ray crystal structures of BPDOB356 with and without its substrate biphenyl 1.6-A˚ resolution for both structures. In both cases, the Fe(II) has five ligands in a square pyramidal configuration: H233 Ne2, H239 Ne2, D386
Od1 and Od2, and a single water molecule. Analysis of the active sites of BPDOB356 and related ROs revealed structural
features that likely contribute to the superior PCB-degrading ability of certain BPDOs. First, the active site cavity readily accommodates biphenyl with minimal conformational rearrangement. Second, M231 was predicted to sterically interfere with binding of some PCBs, and substitution of this residue yielded variants that transform 2,29-dichlorobiphenyl more effectively. Third, in addition to the volume and shape of the active site, residues at the active site entrance also apparently
influence substrate preference. Finally, comparison of the conformation of the active site entrance loop among ROs
provides a basis for a structure-based classification consistent with a phylogeny derived from amino acid sequence
alignments.