Structures of ternary complexes of BphK, a bacterial glutathione S-transferase that reductively dechlorinates polychlorinated biphenyl metabolites.
Prokaryotic glutathione S-transferases are as diverse as their eukaryotic counterparts but are much less well characterized. BphK from Burkholderia xenovorans LB400 consumes two GSH molecules to reductively dehalogenate chlorinated 2-hydroxy-6-oxo-6-phenyl-2,4-dienoates (HOPDAs), inhibitory polychlorinated biphenyl metabolites. Crystallographic structures of two ternary complexes of BphK were solved to a resolution of 2.1A. In the BphK-GSH-HOPDA complex, GSH and HOPDA molecules occupy the G- and H-subsites, respectively. The thiol nucleophile of the GSH molecule is positioned for SN2 attack at carbon 3 of the bound HOPDA. The respective sulfur atoms of conserved Cys-10 and the bound GSH are within 3.0A, consistent with product release and the formation of a mixed disulfide intermediate. In the BphK-(GSH)2 complex, a GSH molecule occupies each of the two subsites. The three sulfur atoms of the two GSH molecules and Cys-10 are aligned suitably for a disulfide exchange reaction that would regenerate the resting enzyme and yield disulfide-linked GSH molecules. A second conserved residue, His-106, is adjacent to the thiols of Cys-10 and the GSH bound to the G-subsite and thus may stabilize a transition state in the disulfide exchange reaction. Overall, the structures support and elaborate a proposed dehalogenation mechanism for BphK and provide insight into the plasticity of the H-subsite.
Tocheva
Elitza I
EI
Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
Fortin
Pascal D
PD
Eltis
Lindsay D
LD
Murphy
Michael E P
ME
D001665:Binding Sites; D019117:Burkholderia / Q000201:enzymology*; D018360:Crystallography, X-Ray; D004220:Disulfides / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000737:chemistry; D005978:Glutathione / Q000737:chemistry; D005982:Glutathione Transferase / Q000737:chemistry*; D011078:Polychlorinated Biphenyls / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary
D004220:Disulfides; D005231:Fatty Acids, Unsaturated; C018971:2-hydroxy-6-oxo-6-phenyl-2,4-hexadienoic acid; D011078:Polychlorinated Biphenyls; D005982:Glutathione Transferase; D005978:Glutathione