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Testing the evolvability of an insect carboxylesterase for the detoxification of synthetic pyrethroid insecticides

Coppin, Christopher W; Jackson, Colin; Sutherland, Tara; Hart, Peter J; Devonshire, Alan L; Russell, Robyn J; Oakeshott , John

Description

Esterases have been implicated in metabolic resistance to synthetic pyrethroids in several insect species but little is yet known of the molecular basis for these effects. In this work modern directed evolution technology was used to test to what extent it is possible to genetically enhance the pyrethroid hydrolytic activity of the E3 carboxylesterase from the blowfly Lucilia cuprina. High throughput screening of a random mutant library with individual stereoisomers of fluorogenic analogues of...[Show more]

dc.contributor.authorCoppin, Christopher W
dc.contributor.authorJackson, Colin
dc.contributor.authorSutherland, Tara
dc.contributor.authorHart, Peter J
dc.contributor.authorDevonshire, Alan L
dc.contributor.authorRussell, Robyn J
dc.contributor.authorOakeshott , John
dc.date.accessioned2015-12-10T22:58:36Z
dc.identifier.issn0965-1748
dc.identifier.urihttp://hdl.handle.net/1885/60930
dc.description.abstractEsterases have been implicated in metabolic resistance to synthetic pyrethroids in several insect species but little is yet known of the molecular basis for these effects. In this work modern directed evolution technology was used to test to what extent it is possible to genetically enhance the pyrethroid hydrolytic activity of the E3 carboxylesterase from the blowfly Lucilia cuprina. High throughput screening of a random mutant library with individual stereoisomers of fluorogenic analogues of two type II pyrethroids identified 17 promising variants that were then also tested with the commercial pyrethroid deltamethrin. Between them, these variants displayed significantly improved activities for all the substrates tested. Amino acid substitutions at ten different residues were clearly implicated in the improvements, although most only enhanced activity for a subset of the stereoisomers. Several new combinations of the most promising amino acid substitutions were then made, and negative epistatic effects were found in most of the combinations, but significant improvements were also found in a minority of them. The best mutant recovered contained three amino acid changes and hydrolysed deltamethrin at more than 100 times the rate of wild-type E3. Structural analysis shows that nine of the ten mutated residues improving pyrethroid or analogue activities cluster in putative substrate binding pockets in the active site, with the three mutations of largest effect all increasing the volume of the acyl pocket.
dc.publisherPergamon-Elsevier Ltd
dc.sourceInsect Biochemistry and Molecular Biology
dc.subjectKeywords: carboxylesterase; cipermethrin; deltamethrin; fenvalerate; insect protein; insecticide; nitrile; pyrethroid; amino acid substitution; animal; article; enzymology; Escherichia coli; fly; genetic recombination; genetics; insecticide resistance; metabolism; Cypermethrin; Deltamethrin; Esterase 3 (E3); Fenvalerate; In vitro evolution
dc.titleTesting the evolvability of an insect carboxylesterase for the detoxification of synthetic pyrethroid insecticides
dc.typeJournal article
local.description.notesImported from ARIES
local.identifier.citationvolume42
dc.date.issued2012
local.identifier.absfor030401 - Biologically Active Molecules
local.identifier.ariespublicationu4005981xPUB574
local.type.statusPublished Version
local.contributor.affiliationCoppin, Christopher W, CSIRO Ecosystem Sciences
local.contributor.affiliationJackson, Colin, College of Physical and Mathematical Sciences, ANU
local.contributor.affiliationSutherland, Tara, CSIRO Ecosystem Sciences
local.contributor.affiliationHart, Peter J, CSIRO Ecosystem Sciences
local.contributor.affiliationDevonshire, Alan L, CSIRO Ecosystem Sciences
local.contributor.affiliationRussell, Robyn J, CSIRO Ecosystem Sciences
local.contributor.affiliationOakeshott , John, CSIRO Ecosystems Science
local.description.embargo2037-12-31
local.bibliographicCitation.issue5
local.bibliographicCitation.startpage343
local.bibliographicCitation.lastpage352
local.identifier.doi10.1016/j.ibmb.2012.01.004
local.identifier.absseo970103 - Expanding Knowledge in the Chemical Sciences
dc.date.updated2016-02-24T10:25:04Z
local.identifier.scopusID2-s2.0-84858071605
local.identifier.thomsonID000302515800004
CollectionsANU Research Publications

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