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Evolution of a microbial acetyltransferase for modification of glyphosate: a novel tolerance strategy
Author(s) -
Siehl Daniel L,
Castle Linda A,
Gorton Rebecca,
Chen Yong Hong,
Bertain Sean,
Cho HyeonJe,
Keenan Robert,
Liu Donglong,
Lassner Michael W
Publication year - 2005
Publication title -
pest management science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.296
H-Index - 125
eISSN - 1526-4998
pISSN - 1526-498X
DOI - 10.1002/ps.1014
Subject(s) - glyphosate , serine , biochemistry , enzyme , transgene , biology , acetylation , genetically modified crops , acetyltransferase , chemistry , microbiology and biotechnology , gene
N ‐Acetylation is a modification of glyphosate that could potentially be used in transgenic crops, given a suitable acetyltransferase. Weak enzymatic activity ( k cat = 5 min −1 , K M = 1 m M ) for N ‐acetylation of glyphosate was discovered in several strains of Bacillus licheniformis (Weigmann) Chester by screening a microbial collection with a mass spectrometric assay. The parental enzyme conferred no tolerance to glyphosate in any host when expressed as a transgene. Eleven iterations of DNA shuffling resulted in a 7000‐fold improvement in catalytic efficiency ( k cat / K M ), sufficient for conferring robust tolerance to field rates of glyphosate in transgenic tobacco and maize. In terms of k cat / K M , the native enzyme exhibited weak activity (4–450% of that with glyphosate) with seven of the common amino acids. Evolution of the enzyme towards an improved k cat / K M for glyphosate resulted in increased activity toward aspartate (40‐fold improved k cat ), but activity with serine and phosphoserine almost completely vanished. No activity was observed among a broad sampling of nucleotides and antibiotics. Improved catalysis with glyphosate coincided with increased thermal stability. Copyright © 2005 Society of Chemical Industry