关键词: 淡色库蚊, 抗药性, 羧酸酯酶, 纯化, 酶动力学, 结合隔离作用

Abstract: In Culex pipiens, resistance to organophosphate insecticides was recognized to be correlated with the increased nonspecific esterase activity. From a dipterexresistant mosquito strain (RD) of Culex pipiens pallens a special protein band was confirmed on SDS/PAGE spectrum. The identified band was not detectable in the susceptible strain(S) or the resmethrin-resistant strain (PY).This band represented a protein of MW 66 kD with pI value 4.6 ,which displayed carboxyesterase activity and accounted for 2.1% of the total extractable proteins. The purification of this protein was performed in turn by phenyl-sepharose (CL-4B), DEAE-cellulose (DE-5L) and FPLC column chromatographic procedures. Enzymological kinetics demonstrated a K_m value of 64.1 mmol/L and a V_max value of 249.8 mmol/(L·mg·min) with α-naphthol acetate as substrate. Dipterex could not inhibit its esterase activity, suggesting the 66 kD protein to be an A-esterase. Comparing with other esterase, it had lower K_m value and V_max value (except higher Vmax value than B-esterase). Further study exhibited that the 66 kD protein might play a role in insect resistance to organophosphate insecticide, mainly through sequestration effect. Nevertheless, the role of hydrolyzing activity at a minor extent could not be excluded.

Key words: Culex pipiens pallens, insecticide resistance, carboxylesterase, purification, enzyme kinetics, sequestration