A SINGLE AMINO-ACID SUBSTITUTION CONVERTS A CARBOXYLESTERASE TO AN ORGANOPHOSPHORUS HYDROLASE AND CONFERS INSECTICIDE RESISTANCE ON A BLOWFLY

Resistance to organophosphorus (OF) insecticides is associated with de creased carboxylesterase activity in several insect species. It has be en proposed that the resistance may be the result of a mutation in a c arboxylesterase that simultaneously reduces its carboxylesterase activ ity and confers an OP hydrolase activity (the ''mutant aliesterase hyp othesis''). In the sheep blowfly, Lucilia cuprina, the association is due to a change in a specific esterase isozyme, E3, which, in resistan t flies, has a null phenotype on gels stained using standard carboxyle sterase substrates. Here we show that an OP-resistant allele of the ge ne that encodes E3 differs at five amino acid replacement sites from a previously described OP-susceptible allele. Knowledge of the structur e of a related enzyme (acetylcholinesterase) suggests that one of thes e substitutions (Gly(137)-->Asp) lies within the active site of the en zyme. The occurrence of this substitution is completely correlated wit h resistance across 15 isogenic strains. In vitro expression of two na tural and two synthetic chimeric alleles shows that the Asp(137) subst itution alone is responsible for both the loss of E3's carboxylesteras e activity and the acquisition of a novel OP hydrolase activity. Model ing of Asp(137) in the homologous position in acetylcholinesterase sug gests that Asp(137) may act as a base to orientate a water molecule in the appropriate position for hydrolysis of the phosphorylated enzyme intermediate.