Discovery and directed evolution of a glyphosate tolerance gene.
The herbicide glyphosate is effectively detoxified by N-acetylation. We screened a collection of microbial isolates and discovered enzymes exhibiting glyphosate N-acetyltransferase (GAT) activity. Kinetic properties of the discovered enzymes were insufficient to confer glyphosate tolerance to transgenic organisms. Eleven iterations of DNA shuffling improved enzyme efficiency by nearly four orders of magnitude from 0.87 mM-1 min-1 to 8320 mM-1 min-1. From the fifth iteration and beyond, GAT enzymes conferred increasing glyphosate tolerance to Escherichia coli, Arabidopsis, tobacco, and maize. Glyphosate acetylation provides an alternative strategy for supporting glyphosate use on crops.
Castle
Linda A
LA
Verdia, Inc. Redwood City, CA 94063, USA. linda.castle@verdiainc.com
Siehl
Daniel L
DL
Gorton
Rebecca
R
Patten
Phillip A
PA
Chen
Yong Hong
YH
Bertain
Sean
S
Cho
Hyeon-Je
HJ
Duck
Nicholas
N
Wong
James
J
Liu
Donglong
D
Lassner
Michael W
MW
D000107:Acetylation; D000123:Acetyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology; D002384:Catalysis; D032621:DNA Shuffling; D019020:Directed Molecular Evolution; D004351:Drug Resistance; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D014644:Genetic Variation; D005998:Glycine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000633:toxicity*; D006540:Herbicides / Q000378:metabolism / Q000633:toxicity*; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D030821:Plants, Genetically Modified / Q000187:drug effects / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D011995:Recombination, Genetic; D014026:Nicotiana / Q000187:drug effects / Q000235:genetics / Q000254:growth & development; D014170:Transformation, Genetic; D003313:Zea mays / Q000187:drug effects / Q000235:genetics / Q000254:growth & development; D000097797:Glyphosate
D000123:Acetyltransferases; D005998:Glycine; D006540:Herbicides; D011994:Recombinant Proteins; D000097797:Glyphosate; C486861:glyphosate N-acetyltransferase