Comparative molecular modeling of Anopheles gambiae CYP6Z1, a mosquito P450 capable of metabolizing DDT

Abstract

One of the challenges faced in malarial control is the acquisition of insecticide resistance that has developed in mosquitoes that are vectors for this disease. Anopheles gambiae, which has been the major mosquito vector of the malaria parasite Plasmodium falciparum in Africa, has over the years developed resistance to insecticides including dieldrin, 1,1-bis(p-chlorophenyl)-2,2,2-trichloroethane (DDT), and pyrethroids. Previous microarray studies using fragments of 230 An. gambiae genes identified five P450 loci, including CYP4C27, CYP4H15, CYP6Z1, CYP6Z2, and CYP12F1, that showed significantly higher expression in the DDT-resistant ZAN/U strain compared with the DDT-susceptible Kisumu strain. To predict whether either of the CYP6Z1 and CYP6Z2 proteins might potentially metabolize DDT, we generated and compared molecular models of these two proteins with and without DDT docked in their catalytic sites. This comparison indicated that, although these two CYP6Z proteins share high sequence identity, their metabolic profiles were likely to differ dramatically from the larger catalytic site of CYP6Z1, potentially involved in DDT metabolism, and the more constrained catalytic site of CYP6Z2, not likely to metabolize DDT. Heterologous expressions of these proteins have corroborated these predictions: only CYP6Z1 is capable of metabolizing DDT. Overlays of these models indicate that slight differences in the backbone of SRS1 and variations of side chains in SRS2 and SRS4 account for the significant differences in their catalytic site volumes and DDT-metabolic capacities. These data identify CYP6Z1 as one important target for inhibitor design aimed at inactivating insecticide-metabolizing P450s in natural populations of this malarial mosquito.cytochrome P450 monooxygenases ͉ insecticides ͉ plant allelochemicals I n 2004, the World Health Organization reported that up to 2.7 million people die of malaria every year with 80-90% of these deaths occurring in Africa (ref. 1 and www.africanfront.com/ AIDS1.php). Many prevention and treatment strategies have been developed to tackle this life-threatening disease from the side of the mosquito vector and that of the human host (2, 3). These range from antimalarial drugs to indoor spraying of insecticides and use of bed nets treated with pyrethroid insecticides. Although these practices have helped reduce human mortality, various issues have emerged with mosquito vectors developing insecticide resistance and parasites developing drug resistance. Both of these significantly reduce the efficacy of current malarial prevention and treatment practices (2, 4-8).The development of insecticide resistance in mosquito vectors is illustrated by Anopheles gambiae, which has been the major mosquito vector of the malaria parasite Plasmodium falciparum in Africa (4, 6). Throughout the years, people have reported Anopheles resistance (albeit low-level resistance) to various insecticides including dieldrin (a cyclodiene-type insecticide), 1,1-bis(pchlorophenyl)-2,2,2-trichloroethane (DDT), and pe...

Esterase-based metabolic resistance to insecticides in heliothine and spodopteran pests

“…One involves mutations in regulatory systems controlling general responses to chemical stresses which upregulate the expression of many gene/enzyme systems with a range of detoxification functions. 15,16) The other involves a population genetic process called 'hitchhiking,' whereby strong positive selection such as that which operates on insecticide resistance mechanisms necessarily entrains large gene frequency changes in other polymorphisms that are genetically linked to those causally involved in resistance. The key point here is that genes encoding detoxifying systems are often arranged in tightly linked clusters in insect genomes.…”

A cis-regulatory sequence driving metabolic insecticide resistance in mosquitoes: Functional characterisation and signatures of selection

“…Confirmatory evidence of a direct role in resistance must come from in vitro metabolism of insecticides with heterologously expressed CYP450, and for a limited number of 4 CYP450s this evidence has been attained (Chiu et al, 2008;Mitchell et al, 2012;Muller et al, 2008;Stevenson et al, 2011). However, whilst gene expression differences coupled with in vitro metabolic activities have confirmed the roles of a number of CYP450s in metabolic resistance to insecticides, to date, the causal mutations that result in this over-expression have remained elusive.…”

Isolation of the monooxygenase complex from Rhipicephalus (Boophilus) microplus – clues to understanding acaricide resistance

“…Four main clades (mitochondrial clade, CYP2, CYP3 and CYP4 clade) exist and in terms of pesticide metabolism the clade that is of interest is the CYP3 clade, a clade which includes the important mammalian CYP3A4 and insect homologues of this enzyme, namely the CYP6 and CYP9 families e.g. CYP6Z1 (Chiu et al, 2008) and CYP6P3 (Muller et al, 2008).…”