COMPARISONS OF KINETIC-PROPERTIES OF ACETYLCHOLINESTERASE PURIFIED FROM AZINPHOSMETHYL-SUSCEPTIBLE AND AZINPHOSMETHYL-RESISTANT STRAINS OF COLORADO-POTATO BEETLE

The affinity (K-m) and hydrolyzing efficiency (V-max) of acetylcholine sterase (AChE, EC 1.1.1.7) purified from a near-isogenic azinphosmethy l-resistant (AZ-R) strain of Colorado potato beetle (CPB) to selected substrates, including acethylthiocholine, acetyl-(beta-methyl) thiocho line, and propionythiocholine, were lower than those of AChE purified from a susceptible (SS) strain. AChE from the SS strain was significan tly inhibited by higher concentrations of acethylthiocholine and acety l-(beta-methyl) thiocholine, whereas AChE from the AZ-R strain was act ivated by higher concentrations of all four substrates examined. AChE from the AZ-R strain was 16- and 1.4-fold less sensitive to inhibition by azinphosmethyl-oxon and dichlorvos, respectively, but was 3.1- and 4.6-fold more sensitive to inhibition by paraoxon and diisopropyl flu orophosphate, respectively, than that from the SS strain. Kinetic stud ies with the selected AChE inhibitors, including eserine, tetramethyla mmonium, tetrapropylammonium, and propidium, indicated that there was no significant difference in inhibition by eserine, tetrapropylammoniu m, and propidium between the two strains. However, AChE from the AZ-R was 1.8-fold less sensitive to inhibition by tetramethylammonium than AChE from the SS strain. Propidium produced a linear mixed-type inhibi tion, whereas the other three inhibitors produced a competitive-type i nhibition in both strains. AChE from the AZ-R strain was also 1.3-fold less sensitive to inhibition by alpha-chaconine, but was 1.7- and 1.8 -fold more sensitive to alpha-solanine and tomatine, respectively, tha n AChE from the: SS strain. These findings support our original conten tion that this aspect of overall azinphosmethyl resistance in CPB was due to a modified form of AChE that functions as a site insensitivity resistance mechanism. Modifications of AChE obtained from the AZ-R str ain may occur both in the catalytic center and at the peripheral anion ic site or in a single region where it modifies the binding of these l igands to both sites, (C) 1995 Academic Press, Inc.