Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.491091
Title: Plant carboxylesterases involved in pesticide hydrolysis
Author: Gershater, Markus Christian
ISNI:       0000 0001 3495 3342
Awarding Body: Durham University
Current Institution: Durham University
Date of Award: 2006
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Abstract:
Many herbicides are applied in the form of carboxylic esters to increase their hydrophobicity and hence aid their passage through the waxy cuticle. Hydrolysis in planta of these pro-herbicide esters releases the active acid or alcohol and the rate of this cleavage can be a factor in determining herbicide selectivity. Protein extracts from 13 crop and weed species were assayed for carboxylesterase activity toward multiple xenobiotic and pesticide ester substrates, including 2,4-D-methyl, aryloxyphenoxypropionate esters and p-nitrophenyl esters. A diversity of activities was exhibited by the different species, with Arabidopsis thaliana extracts showing high hydrolytic activity toward several xenobiotic esters, particularly 2,4-D- methyl.The major 2,4-D-methyl hydrolysing enzyme in arabidopsis cell cultures was purified through three rounds of chromatography, then selectively labelled with a biotinylated fluorophosphonate probe (FP-biotin). Following streptavidin affinity purification, the labelled protein was identified by proteomics as the previously uncharacterised serine hydrolase AtCXE12. Recombinant AtCXE12 was subsequently confirmed to effectively hydrolyse 2,4-D-methyl.A T-DNA insertion knockout line that did not express AtCXE12 was identified and characterised. Protein from the knockout plants did not contain AtCXE12 and was found to have a reduced rate of 2,4-D-methyl hydrolysis compared to wild-type plant extracts. This translated into a higher tolerance of 2,4-D-methyl in young atcxel2 plants, due to a lower rate of bioactivation of the pro-herbicide.The fluorophosphonate-based chemical probe was subsequently used to identify other major serine hydrolases in arabidopsis. AtCXE12 and three previously uncharacterised hydrolases were identified, each belonging to a distinct enzyme family.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.491091  DOI: Not available
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