Identification of an opd (organophosphate degradation) gene in an Agrobacterium isolate.
We isolated a bacterial strain, Agrobacterium radiobacter P230, which can hydrolyze a wide range of organophosphate (OP) insecticides. A gene encoding a protein involved in OP hydrolysis was cloned from A. radiobacter P230 and sequenced. This gene (called opdA) had sequence similarity to opd, a gene previously shown to encode an OP-hydrolyzing enzyme in Flavobacterium sp. strain ATCC 27551 and Brevundimonas diminuta MG. Insertional mutation of the opdA gene produced a strain lacking the ability to hydrolyze OPs, suggesting that this is the only gene encoding an OP-hydrolyzing enzyme in A. radiobacter P230. The OPH and OpdA proteins, encoded by opd and opdA, respectively, were overexpressed and purified as maltose-binding proteins, and the maltose-binding protein moiety was cleaved and removed. Neither protein was able to hydrolyze the aliphatic OP malathion. The kinetics of the two proteins for diethyl OPs were comparable. For dimethyl OPs, OpdA had a higher k(cat) than OPH. It was also capable of hydrolyzing the dimethyl OPs phosmet and fenthion, which were not hydrolyzed at detectable levels by OPH.
Horne
Irene
I
CSIRO Entomology, GPO Box 1700, Canberra, ACT 2601, Australia. irene.horne@ento.csiro.au
Sutherland
Tara D
TD
Harcourt
Rebecca L
RL
Russell
Robyn J
RJ
Oakeshott
John G
JG
D000595:Amino Acid Sequence; D043303:Aryldialkylphosphatase; D003001:Cloning, Molecular; D003372:Coumaphos / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D004950:Esterases / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism; D012231:Rhizobium / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D004950:Esterases; C045290:phosphorylphosphatase; D010744:Phosphoric Monoester Hydrolases; D043303:Aryldialkylphosphatase; C007130:coroxon; D003372:Coumaphos