The bacterial YbaK protein is a Cys-tRNAPro and Cys-tRNA Cys deacylase

Abstract

Bacterial prolyl-tRNA synthetases and some smaller paralogs, YbaK and ProX, can hydrolyze misacylated Cys-tRNAPro or Ala-tRNAPro. To assess the significance of this quality control editing reaction in vivo, we tested Escherichia coli ybaK for its ability to suppress the E. coli thymidylate synthase thyA:146CCA missense mutant strain, which requires Cys-tRNA Pro for growth in the absence of thymine. Missense suppression was observed in a ybaK deletion background, suggesting that YbaK functions as a Cys-tRNAPro deacylase in vivo. In vitro studies with the full set of 20 E. coli aminoacyl-tRNAs revealed that the Haemophilus influenzae and E. coli YbaK proteins are moderately general aminoacyl-tRNA deacylases that preferentially hydrolyze Cys-tRNAPro and Cys-tRNACys and are also weak deacylases that cleave Gly-tRNA, Ala-tRNA, Ser-tRNA, Pro-tRNA, and Met-tRNA. The ProX protein acted as an aminoacyl-tRNA deacylase that cleaves preferentially Ala-tRNA and Gly-tRNA. The potential of H. influenzae YbaK to hydrolyze in vivo correctly charged Cys-tRNACys was tested in E. coli strain X2913 (ybaK+). Overexpression of H. influenzae ybaK decreased the in vivo ratio of Cys-tRNACys to tRNACys from 65 to 35% and reduced the growth rate of strain X2913 by 30% in LB medium. These data suggest that YbaK-mediated hydrolysis of aminoacyl-tRNA can influence cell growth. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.