Structural basis for the enantiospecificities of R‐ and S‐specific phenoxypropionate/α‐ketoglutarate dioxygenases

Abstract

Abstract(R)-and (S)-dichlorprop/a-ketoglutarate dioxygenases (RdpA and SdpA) catalyze the oxidative cleavage of 2-(2,4-dichlorophenoxy)propanoic acid (dichlorprop) and 2-(4-chloro-2-methyl-phenoxy)propanoic acid (mecoprop) to form pyruvate plus the corresponding phenol concurrent with the conversion of a-ketoglutarate (aKG) to succinate plus CO 2 . RdpA and SdpA are strictly enantiospecific, converting only the (R) or the (S) enantiomer, respectively. Homology models were generated for both enzymes on the basis of the structure of the related enzyme TauD (PDB code 1OS7). Docking was used to predict the orientation of the appropriate mecoprop enantiomer in each protein, and the predictions were tested by characterizing the activities of site-directed variants of the enzymes. Mutant proteins that changed at residues predicted to interact with (R)-or (S)-mecoprop exhibited significantly reduced activity, often accompanied by increased K m values, consistent with roles for these residues in substrate binding. Four of the designed SdpA variants were (slightly) active with (R)-mecoprop. The results of the kinetic investigations are consistent with the identification of key interactions in the structural models and demonstrate that enantiospecificity is coordinated by the interactions of a number of residues in RdpA and SdpA. Most significantly, residues Phe171 in RdpA and Glu69 in SdpA apparently act by hindering the binding of the wrong enantiomer more than the correct one, as judged by the observed decreases in K m when these side chains are replaced by Ala.

Enantioselective Dechlorination of Polychlorinated Biphenyls in Dehalococcoides mccartyi CG1

“…The difference in enantioselectivity between the aerobic organohalide-degrading bacteria and the anaerobic organohalide-respiring bacteria might be due to their distinct enzymatic processes. Previous studies on the enantioselective degradation of mecoprop by Sphingomonas herbicidovorans MH implied that the enantioselectivity was coordinated by the interactions of multiple residues in RdpA and SdpA with the substrate (37). For the enantiomer-specific binding of mecoprop to RdpA and SdpA, the active sites could be expected to interact with the carboxylic acid, the ether oxygen atom, and the methyl group by several residues (37).…”

“…Previous studies on the enantioselective degradation of mecoprop by Sphingomonas herbicidovorans MH implied that the enantioselectivity was coordinated by the interactions of multiple residues in RdpA and SdpA with the substrate (37). For the enantiomer-specific binding of mecoprop to RdpA and SdpA, the active sites could be expected to interact with the carboxylic acid, the ether oxygen atom, and the methyl group by several residues (37). In contrast, the organohaliderespiring bacteria employ periplasmic, outward-oriented, membrane-bound RDases to catalyze halogen removal from organohalides without destroying the corresponding backbone carbon structure (e.g., the biphenyl in PCBs) (38).…”

Establishment of Bacterial Herbicide Degraders in a Rapid Sand Filter for Bioremediation of Phenoxypropionate-Polluted Groundwater

“…Phenoxypropionates are chemical enantiomers that exist as both R and S enantiomers. Different bacterial enzymes, namely, the (R)-and (S)-dichlorprop ␣-ketoglutarate dioxygenases, encoded by the rdpA and sdpA genes, respectively, are responsible for the first step in their degradation (6). Under aerobic conditions, the herbicides are subject to rapid biodegradation, and half-lives in topsoils are usually measured in days or weeks (7,8).…”

Molecular basis for enantioselective herbicide degradation imparted by aryloxyalkanoate dioxygenases in transgenic plants

“…The carboxylic acid of (R)-dichlorprop is within hydrogen-bonding distance of both the side chain and backbone nitrogen of Ser114, while the p-chloro atom is within halogen-bonding distance of Asp108 and Asp109. Prior modeling studies based on TauD predicted binding of the ligand in an extended orientation (53), while our structural data show that (R)-dichlorprop binds in a bent conformation that directs the phenoxy ring toward a solvent-exposed opening.…”