Discovery of Escherichia coli methionyl-tRNA synthetase mutants for efficient labeling of proteins with azidonorleucine in vivo.

ABSTRACT: Incorporation of noncanonical amino acids into cellular proteins often requires engineering new aminoacyl-tRNA synthetase activity into the cell. A screening strategy that relies on cell-surface display of reactive amino acid side-chains was used to identify a diverse set of methionyl-tRNA synthetase (MetRS) mutants that allow efficient incorporation of the methionine (Met) analog azidonorleucine (Anl). We demonstrate that the extent of cell-surface labeling in vivo is a good indicator of the rate of Anl activation by the MetRS variant harbored by the cell. By screening at low Anl concentrations in Met-supplemented media, MetRS variants with improved activities toward Anl and better discrimination against Met were identified.

SUBMITTER: Tanrikulu IC 

PROVIDER: S-EPMC2741243 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature