A cluster of esterase genes on chromosome 3R ofDrosophila melanogaster includes homologues of esterase genes conferring insecticide resistance inLucilia cuprina

Spackman, M.; Oakeshott, John G.; Smyth, Kerrie-Ann; Medveczky, Kerrie M.; Russell, Robyn J.

doi:10.1007/bf00557238

Cited by 27 publications

(24 citation statements)

References 40 publications

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“…2 A and B). α-CBEs have long been known to be associated with the microsomal fraction (17). However, it was unknown how this association was maintained at a structural level.…”

Section: Resultsmentioning

confidence: 99%

Structure and function of an insect α-carboxylesterase (αEsterase7) associated with insecticide resistance

Jackson

Liu

Carr

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Insect carboxylesterases from the αEsterase gene cluster, such as αE7 (also known as E3) from the Australian sheep blowfly Lucilia cuprina (LcαE7), play an important physiological role in lipid metabolism and are implicated in the detoxification of organophosphate (OP) insecticides. Despite the importance of OPs to agriculture and the spread of insect-borne diseases, the molecular basis for the ability of α-carboxylesterases to confer OP resistance to insects is poorly understood. In this work, we used laboratory evolution to increase the thermal stability of LcαE7, allowing its overexpression in Escherichia coli and structure determination. The crystal structure reveals a canonical α/β-hydrolase fold that is very similar to the primary target of OPs (acetylcholinesterase) and a unique N-terminal α-helix that serves as a membrane anchor. Soaking of LcαE7 crystals in OPs led to the capture of a crystallographic snapshot of LcαE7 in its phosphorylated state, which allowed comparison with acetylcholinesterase and rationalization of its ability to protect insects against the effects of OPs. Finally, inspection of the active site of LcαE7 reveals an asymmetric and hydrophobic substrate binding cavity that is well-suited to fatty acid methyl esters, which are hydrolyzed by the enzyme with specificity constants (∼10 6 M −1 s −1 ) indicative of a natural substrate.protein engineering | directed evolution | ali-esterase T he demand for greater productivity from agriculture and the avoidance of insect-borne diseases has made efficient control of insect pests increasingly important; in 2007, the world market for insecticides was estimated to be in the order of $11 billion US (1). However, the effectiveness of insecticides is decreasing, with over 500 documented instances of insecticide resistance (2). The development of resistance to organophosphate (OP) (Fig. 1A) pesticides through the action of α-carboxylesterases (α-CBEs) has been documented in many species (3), including the Australian sheep blowfly Lucilia cuprina (4). OPs inhibit acetylcholinesterase (AChE) at cholinergic synapses in the central and peripheral nervous systems, which leads to interminable nerve signal transduction and death (5). However, α-CBEs, such as αE7 in L. cuprina (LcαE7), confer a significant protective effect on insects owing to their ability to bind and slowly hydrolyze OPs (6). It has also been shown that this CBE-mediated OP resistance can be increased through G137D and W251L point mutations (4, 7).Insect α-CBEs, including LcαE7, are thought to play important physiological roles in lipid and xenobiotic metabolism (6,8). They are one of the most abundant protein families in insects, with between 20 and 40 active CBEs predicted to be expressed by the fruit fly Drosphila melanogaster (9, 10). Although the physiological substrates of insect α-CBEs are unknown, recent MS results have shown that D. melanogaster αE7 is expressed in the fat body lipid droplet proteome (11), implying a role in lipid or cholesterol metabolism. This work has been ext...

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“…2 A and B). α-CBEs have long been known to be associated with the microsomal fraction (17). However, it was unknown how this association was maintained at a structural level.…”

Section: Resultsmentioning

confidence: 99%

Structure and function of an insect α-carboxylesterase (αEsterase7) associated with insecticide resistance

Jackson

Liu

Carr

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

120

123

View full text Add to dashboard Cite

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“…Carboxyl-esterases are related to many roles: resistance to organophosphorus (OP) (Newcomb et al, , 1997aCampbell et al, 1998a, b;Claudianos, Russell & Oakeshott, 1999), basis of many shared physiological and biochemical characteristics (Spackman et al, 1994;Parker et al, 1996;Campbell et al, 1997) and digestive activity (Campbell et al, 2003). The inheritance pattern of the Est-3 locus was shown to be compatible to monogenic Mendelian segregation of its four alleles.…”

Section: Discussionmentioning

confidence: 95%

“…In insects, they are related to several metabolic processes, such as food digestion, degradation of insecticides, pheromones and juvenile hormone hydrolysis (Spackman et al, 1994;Campbell et al, 2003). They also seem to be related to sexual activity (Richmond & Senior, 1991;Korochkin, 1995;Tamarina, Ludwig & Richmond, 1997;Oakeshott et al, 2001).…”

Section: Introductionmentioning

confidence: 94%

Characterization of Esterases in a Brazilian Population of Zaprionus Indianus (Diptera: Drosophilidae)

2006

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The aim of this study was to characterize esterases in Zaprionus indianus, a drosophilid recently introduced into Brazil. A further aim was study the variation of activity of esterases in the presence of inhibitors and their expression according to sex, sexual activity and age of individual flies. Polymorphisms were detected in two esterase loci (Est-2 and Est-3) and monomorphisms in four others (Est-1, Est-4, Est-5 and Est-6). Biochemical tests using alpha- and beta-naphthyl acetate and the inhibitors malathion, eserine sulphate and PMSF allowed us to classify EST-2 and EST-5 as beta-esterases, both carboxyl-esterases, and EST-1, EST-3, EST-4 and EST-6 as alpha-esterases. EST-1 and EST-3 were classified as carboxyl-esterases and EST-4 and EST-6 as cholinesterases. EST-5 activity was more pronounced in males and EST-2 was restricted to them or to recently copulated females. EST-4, rarely detected, was not characterized. Based on their biochemical characteristics possible roles for these enzymes are suggested.

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“…1A) corresponds to the gene product of the a-Esterase-7 (a-Est7) gene (also called CG1112; www.flybase.org). The protein product of a-Est7, a-Esterase-7 (a-EST7) is a membrane-associated a-Esterase with preference for substrates with shorter acid side chain length (Spackman et al, 1994;Parker et al, 1996). Inhibitor sensitivity studies classify a-EST7 as subclass I CE (Spackman et al, 1994).…”

Section: Functional Proteomics Of Drosophila Fat Body Esterasesmentioning

confidence: 99%

Functional fat body proteomics and gene targeting reveal in vivo functions of Drosophila melanogaster α-Esterase-7

Birner‐Gruenberger

Bickmeyer

Lange

et al. 2012

Insect Biochemistry and Molecular Biology

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Carboxylesterases constitute a large enzyme family in insects, which is involved in diverse functions such as xenobiotic detoxification, lipid metabolism and reproduction. Phylogenetically, many insect carboxylesterases are represented by multienzyme clades, which are encoded by evolutionarily ancient gene clusters such as the α-Esterase cluster. Much in contrast to the vital importance attributed to carboxylesterases in general, the in vivo function of individual α-Esterase genes is largely unknown. This study employs a functional proteomics approach to identify esterolytic enzymes of the vinegar fly Drosophila melanogaster fat body. One of the fat body carboxylesterases, α-Esterase-7, was selected for mutational analysis by gene targeting to generate a deletion mutant fly. Phenotypic characterization of α-Esterase-7 null mutants and transgenic flies, which overexpress a chimeric α-Esterase-7:EGFP gene, reveals important functions of α-Esterase-7 in insecticide tolerance, lipid metabolism and lifespan control. The presented first deletion mutant of any α-Esterase in the model insect D. melanogaster generated by gene targeting not only provides experimental evidence for the endogenous functions of this gene family. It also offers an entry point for in vivo structure-function analyses of α-Esterase-7, which is of central importance for naturally occurring insecticide resistance in wild populations of various dipteran insect species.

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A cluster of esterase genes on chromosome 3R ofDrosophila melanogaster includes homologues of esterase genes conferring insecticide resistance inLucilia cuprina

Cited by 27 publications

References 40 publications

Structure and function of an insect α-carboxylesterase (αEsterase7) associated with insecticide resistance

Structure and function of an insect α-carboxylesterase (αEsterase7) associated with insecticide resistance

Characterization of Esterases in a Brazilian Population of Zaprionus Indianus (Diptera: Drosophilidae)

Functional fat body proteomics and gene targeting reveal in vivo functions of Drosophila melanogaster α-Esterase-7

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