(Below N is a link to NCBI taxonomic web page and E link to ESTHER at designed phylum.) > cellular organisms: NE > Eukaryota: NE > Opisthokonta: NE > Metazoa: NE > Eumetazoa: NE > Bilateria: NE > Protostomia: NE > Ecdysozoa: NE > Panarthropoda: NE > Arthropoda: NE > Mandibulata: NE > Pancrustacea: NE > Hexapoda: NE > Insecta: NE > Dicondylia: NE > Pterygota: NE > Neoptera: NE > Holometabola: NE > Diptera: NE > Brachycera: NE > Muscomorpha: NE > Eremoneura: NE > Cyclorrhapha: NE > Schizophora: NE > Acalyptratae: NE > Ephydroidea: NE > Drosophilidae: NE > Drosophilinae: NE > Drosophilini: NE > Drosophila [fruit fly, genus]: NE > Sophophora: NE > melanogaster group: NE > melanogaster subgroup: NE > Drosophila melanogaster: NE
LegendThis sequence has been compared to family alignement (MSA) red => minority aminoacid blue => majority aminoacid color intensity => conservation rate title => sequence position(MSA position)aminoacid rate Catalytic site Catalytic site in the MSA TIEHKVQQYRQSTNETVVADTEYGQVRGIKRLSLYDVPYFSFEGIPYAQP PVGELRFKAPQRPIPWEGVRDCSQPKDKAVQVQFVFDKVEGSEDCLYLNV YTNNVKPDKARPVMVWIHGGGFIIGEANREWYGPDYFMKEDVVLVTIQYR LGALGFMSLKSPELNVPGNAGLKDQVLALKWIKNNCASFGGDPNCITVFG ESAGGASTHYMMLTDQTQGLFHRGILQSGSAICPWAYNGDITHNPYRIAK LVGYKGEDNDKDVLEFLQNVKAKDLIRVEENVLTLEERMNKIMFRFGPSL EPFSTPECVISKPPKEMMKTAWSNSIPMFIGNTSYEGLLWVPEVKLMPQV LQQLDAGTPFIPKELLATEPSKEKLDSWSAQIRDVHRTGSESTPDNYMDL CSIYYFVFPALRVVHSRHAYAAGAPVYFYRYDFDSEELIFPYRIMRMGRG VKGVSHADDLSYQFSSLLARRLPKESREYRNIERTVGIWTQFAATGNPYS EKINGMDTLTIDPVRKSDEVIKCLNISDDLKFIDLPEWPKLKVWESLYDD NKDLLF
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 a-Esterase cluster. Much in contrast to the vital importance attributed to carboxylesterases in general, the in vivo function of individual a-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, a-Esterase-7, was selected for mutational analysis by gene targeting to generate a deletion mutant fly. Phenotypic characterization of a-Esterase-7 null mutants and transgenic flies, which overexpress a chimeric a-Esterase-7:EGFP gene, reveals important functions of a-Esterase-7 in insecticide tolerance, lipid metabolism and lifespan control. The presented first deletion mutant of any a-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 a-Esterase-7, which is of central importance for naturally occurring insecticide resistance in wild populations of various dipteran insect species.
Title: Hydrolysis of pyrethroids by carboxylesterases from Lucilia cuprina and Drosophila melanogaster with active sites modified by in vitro mutagenesis Heidari R, Devonshire AL, Campbell BE, Dorrian SJ, Oakeshott JG, Russell RJ Ref: Insect Biochemistry & Molecular Biology, 35:597, 2005 : PubMed
The cloned genes encoding carboxylesterase E3 in the blowfly Lucilia cuprina and its orthologue in Drosophila melanogaster were expressed in Sf9 cells transfected with recombinant baculovirus. Resistance of L. cuprina to organophosphorus insecticides is due to mutations in the E3 gene that enhance the enzyme's ability to hydrolyse insecticides. Previous in vitro mutagenesis and expression of these modifications (G137D, in the oxyanion hole and W251L, in the acyl pocket) have confirmed their functional significance. We have systematically substituted these and nearby amino acids by others expected to affect the hydrolysis of pyrethroid insecticides. Most mutations of G137 markedly decreased pyrethroid hydrolysis. W251L was the most effective of five substitutions at this position. It increased activity with trans permethrin 10-fold, and the more insecticidal cis permethrin >130-fold, thereby decreasing the trans:cis hydrolysis ratio to only 2, compared with >25 in the wild-type enzyme. Other mutations near the bottom of the catalytic cleft generally enhanced pyrethroid hydrolysis, the most effective being F309L, also in the presumptive acyl binding pocket, which enhanced trans permethrin hydrolysis even more than W251L. In these assays with racemic 1RS cis and 1RS trans permethrin, two phases were apparent, one being much faster suggesting preferential hydrolysis of one enantiomer in each pair as found previously with other esterases. Complementary assays with individual enantiomers of deltamethrin and the dibromo analogue of cis permethrin showed that the wild type and most mutants showed a marked preference for the least insecticidal 1S configuration, but this was reversed by the F309L substitution. The W251L/F309L double mutant was best overall in hydrolysing the most insecticidal 1R cis isomers. The results are discussed in relation to likely steric effects on enzyme-substrate interactions, cross-resistance between pyrethroids and malathion, and the potential for bioremediation of pyrethroid residues.
Title: Duplication and divergence of the genes of the alpha-esterase cluster of Drosophila melanogaster Robin C, Russell RJ, Medveczky KM, Oakeshott JG Ref: Journal of Molecular Evolution, 43:241, 1996 : PubMed
The alpha-esterase cluster of D. melanogaster contains 11 esterase genes dispersed over 60 kb. Embedded in the cluster are two unrelated open reading frames that have sequence similarity with genes encoding ubiquitin-conjugating enzyme and tropomyosin. The esterase amino acid sequences show 37-66% identity with one another and all but one have all the motifs characteristic of functional members of the carboxyl/cholinesterase multigene family. The exception has several frameshift mutations and appears to be a pseudogene. Patterns of amino acid differences among cluster members in relation to generic models of carboxyl/cholinesterase protein structure are broadly similar to those among other carboxyl/cholinesterases sequenced to date. However the alpha-esterases differ from most other members of the family in: their lack of a signal peptide; the lack of conservation in cysteines involved in disulfide bridges; and in four indels, two of which occur in or adjacent to regions that align with proposed substrate-binding sites of other carboxyl/cholinesterases. Phylogenetic analyses clearly identify three simple gene duplication events within the cluster. The most recent event involved the pseudogene which is located in an intron of another esterase gene. However, relative rate tests suggest that the pseudogene remained functional after the duplication event and has become inactive relatively recently. The distribution of indels also suggests a deeper node in the gene phylogeny that separates six genes at the two ends of the cluster from a block of five in the middle.
Drosophila melanogaster (Fruit fly) acetylcholinesterase H subunit