Permethrin Carboxylesterase Functions as Nonspecific Sequestration Proteins in the Hemolymph of Colorado Potato Beetle

Abstract

The inhibition profiles of the pI4.8 and 4.5 carboxylesterases from the permethrin-resistant strain of Colorado potato beetle were evaluated for a variety of inhibitors and insecticides. Both carboxylesterases were determined to be serine-hydroxyl-type esterases with different inhibition specificities toward phenylmethylsulfonyl fluoride, a serine-hydroxyl proteinase inhibitor, andpara-hydroxylmercuribenzoate, a cysteine-sulfhydryl group arylesterase inhibitor. Thepara-hydroxylmercuribenzoate-binding site appears separate from the catalytic binding site as determined byp-chloromercuribenzoate-agarose affinity chromatography of the purified pI4.5–4.8 carboxylesterases. The pI4.8 and 4.5 carboxylesterases, however, were not significantly different in their sensitivity to various insecticides. The levels of inhibition achieved by the insecticides and eserine showed a positive correlation with the hydrophobicity of the compounds, suggesting that the pI4.8 and 4.5 carboxylesterases can interact with a variety of hydrophobic compounds through nonspecific hydrophobic site(s). The kinetics of inhibition of the pI4.5–4.8 carboxylesterases elicited by permethrin and DDT are most similar to a mixed-noncompetitive type and a noncompetitive type of inhibition, respectively. Analysis of these kinetic differences indicates the presence of hydrophobic catalytic site(s) as well as hydrophobic noncatalytic site(s) that are available for the binding of esteratic (e.g., permethrin) and nonesteratic (e.g., DDT) hydrophobic insecticides, respectively. Along with a low level of permethrin hydrolysis, the hydrophobic binding nature of the pI4.5–4.8 carboxylesterases suggests that resistance to permethrin is mainly conferred by sequestration. Sequestration of insecticides by the pI4.5–4.8 carboxylesterases appears to be nonspecific and associated with the cross-resistance of the PE-R strain to other hydrophobic insecticides, such as other pyrethroids, DDT, and abamectin.