Karunaratne S H, Hemingway J, Jayawardena K G, Dassanayaka V, Vaughan A
Department of Pure and Applied Biology, University of Wales Cardiff, Wales.
J Biol Chem. 1995 Dec 29;270(52):31124-8. doi: 10.1074/jbc.270.52.31124.
Two non-amplified esterases were purified from the insecticide-susceptible Pel SS strain of Culex quinquefasciatus. These were the two major esterase activity peaks in this strain. The two corresponding amplified carboxylesterases, Est alpha 2 and Est beta 2, involved in organophosphate sequestration were purified from two resistant C. quinquefasciatus strains. The Pel SS esterases were significantly less reactive with the organophosphates than those from the resistant strains. One of the Pel SS esterases was electrophoretically identical to amplified Culex Est beta 1. However, it differed kinetically, and in its nucleotide and predicted amino acid sequences from the two characterized amplified Est beta 1s, it is classified as Est beta 1(3). Restriction fragment analysis suggested Pel SS has only one Est alpha and one Est beta gene, while the resistant Pel RR has both amplified and non-amplified forms of Est alpha and Est beta. The EcoRI fragments for both Pel SS esterases were distinct from those of the amplified Est alpha 2(1), Est beta 2(1), or Est beta 1(1&2). An esterase with the same size EcoRI fragment as Est beta 1(3) was also present in Pel RR. This and restriction enzyme fragment analysis of C. quinquefasciatus field populations suggest that variability of the susceptible alleles may be lower than previously suggested. A non-amplified Est alpha with a unique EcoRI band was present in Pel RR. The previous esterase purification procedures may not have separated these amplified and non-amplified alleles. Hence, the small differences between the purified esterases from resistant strains may reflect mixtures of identical amplified alleles with different non-amplified alleles, which have significantly different k alpha values.
从对杀虫剂敏感的致倦库蚊Pel SS品系中纯化出两种非扩增酯酶。它们是该品系中的两个主要酯酶活性峰。从两个抗性致倦库蚊品系中纯化出了与有机磷螯合相关的两种相应的扩增羧酸酯酶，即Est alpha 2和Est beta 2。与抗性品系的酯酶相比，Pel SS酯酶对有机磷的反应活性明显较低。Pel SS酯酶之一在电泳上与扩增的库蚊Est beta 1相同。然而，它在动力学上以及核苷酸和预测的氨基酸序列方面与两个已鉴定的扩增Est beta 1不同，因此被归类为Est beta 1(3)。限制性片段分析表明，Pel SS只有一个Est alpha基因和一个Est beta基因，而抗性品系Pel RR同时具有扩增型和非扩增型的Est alpha和Est beta。Pel SS两种酯酶的EcoRI片段与扩增的Est alpha 2(1)、Est beta 2(1)或Est beta 1(1&2)的片段不同。Pel RR中也存在一种与Est beta 1(3)具有相同大小EcoRI片段的酯酶。这以及对致倦库蚊野外种群的限制性酶切片段分析表明，敏感等位基因的变异性可能低于先前的推测。Pel RR中存在一个具有独特EcoRI条带的非扩增Est alpha。先前的酯酶纯化程序可能没有分离出这些扩增和非扩增的等位基因。因此，抗性品系纯化酯酶之间的微小差异可能反映了相同扩增等位基因与不同非扩增等位基因的混合物，它们具有显著不同的k alpha值。
