Discovery and Improvement of gat Genes

To discover an enzyme capable of acetylating glyphosate, we searched our microbial

collection for an organism that could carry out the reaction. Since bacilli produce a
wide variety of enzymes involved in secondary metabolism, we focused
on Bacillus and related genera isolated from non-extreme environments. We
developed a sensitive, high throughput (HTP) MS-based assay capable of detecting N-
acetyl glyphosate in crude cell extracts. When incubated with glyphosate and acetyl
CoA, strains ofBacillus licheniformis, a common saprophytic bacterium, catalyzed
reproducible accumulation of the acetylated herbicide. To isolate the gene encoding
GAT, recombinant E. coli, expressing genomic DNA fragments from B.
licheniformis, were assayed by the MS method. N-acetyltransferase genes isolated
from these strains are 93% identical. GAT enzymes are ~17 kDa, are most active at