Non-specific activities of the major herbicide-resistance gene BAR

Christ, Bastien; Hochstrasser, Ramon; Guyer, Luzia; Francisco, Rita; Aubry, Sylvain; Hörtensteiner, Stefan; Weng, Jing-Ke

doi:10.1038/s41477-017-0061-1

Non-specific activities of the major herbicide-resistance gene BAR

Journal Article · Mon Nov 27 04:00:00 EST 2017 · Nature Plants (Online)

DOI:https://doi.org/10.1038/s41477-017-0061-1· OSTI ID:1430306

Christ, Bastien ^[1]; Hochstrasser, Ramon ^[2]; Guyer, Luzia ^[2]; Francisco, Rita ^[2]; ^[2]; ^[2]; ^[3]

Whitehead Inst. for Biomedical Research, Cambridge, MA (United States)
Univ. of Zurich (Switzerland)
Whitehead Inst. for Biomedical Research, Cambridge, MA (United States); Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States)

Bialaphos resistance (BAR) and phosphinothricin acetyltransferase (PAT) genes, which convey resistance to the broad-spectrum herbicide phosphinothricin (also known as glufosinate) via N-acetylation, have been globally used in basic plant research and genetically engineered crops. Although early in vitro enzyme assays showed that recombinant BAR and PAT exhibit substrate preference toward phosphinothricin over the 20 proteinogenic amino acids, indirect effects of BAR-containing transgenes in planta, including modified amino acid levels, have been seen but without the identification of their direct causes. Combining metabolomics, plant genetics and biochemical approaches, we show that transgenic BAR indeed converts two plant endogenous amino acids, aminoadipate and tryptophan, to their respective N-acetylated products in several plant species. We report the crystal structures of BAR, and further delineate structural basis for its substrate selectivity and catalytic mechanism. Through structure-guided protein engineering, we generated several BAR variants that display significantly reduced non-specific activities compared with its wild-type counterpart in vivo. Here, the transgenic expression of enzymes can result in unintended off-target metabolism arising from enzyme promiscuity. Understanding such phenomena at the mechanistic level can facilitate the design of maximally insulated systems featuring heterologously expressed enzymes.

View Accepted Manuscript (DOE)

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22). Scientific User Facilities Division; National Inst. of General Medical Sciences; National Inst of Health; NIH-ORIP HEI; Swiss National Science Foundation

Grant/Contract Number:: AC02-06CH11357

OSTI ID:: 1430306

Journal Information:: Nature Plants (Online), Journal Name: Nature Plants (Online) Journal Issue: 12 Vol. 3; ISSN 2055-0278

Publisher:: Nature Publishing GroupCopyright Statement

Country of Publication:: United States

Language:: ENGLISH

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