5T7E

Crystal structure of Streptomyces hygroscopicus Bialaphos Resistance (BAR) protein in complex with Coenzyme A and L-phosphinothricin

Summary for 5T7E

• Entry DOI: 10.2210/pdb5t7e/pdb
• Related: 5T7D
• Descriptor: Phosphinothricin N-acetyltransferase, COENZYME A, PHOSPHINOTHRICIN, ... (5 entities in total)
• Functional Keywords: phosphinothricin-deactivating activity gcn5-related n-acetyltransferases, transferase
• Biological source: Streptomyces hygroscopicus
• Total number of polymer chains: 4
• Total formula weight: 89531.64

Authors

Christ, B.,Weng, J.K. (deposition date: 2016-09-04, release date: 2017-06-07, Last modification date: 2024-05-22)

Primary citation

Christ, B.,Hochstrasser, R.,Guyer, L.,Francisco, R.,Aubry, S.,Hortensteiner, S.,Weng, J.K.
Non-specific activities of the major herbicide-resistance gene BAR.
Nat Plants, 3:937-945, 2017

PubMed Abstract: 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. 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.

PubMed: 29180815
DOI: 10.1038/s41477-017-0061-1

Experimental method

X-RAY DIFFRACTION (1.8 Å)