Anticodon‐Binding Domain Mutations of the Bacillus subtilis Non‐discriminating Aspartyl‐tRNA Synthetase to Increase tRNA Specificity

Abstract

The fidelity of the genetic code is etablished by tRNA synthetases attaching an amino acid to the correct tRNA isoacceptor. The majority of prokaryotes lack an asparaginyl‐tRNA synthetase to ligate asparagine to tRNAAsn. Instead, these organisms possess a non‐discriminating aspartyl‐tRNA synthetase (ND‐AspRS), an AspRS with relaxed substrate specificity, that can ligate aspartate to either tRNAAsn or tRNAAsp. An amidotransferase enzyme, GatCAB, then amidates Asp‐tRNAAsn to form Asn‐tRNAAsn. In many but not all organisms encoding an asparaginyl‐tRNA synthetase to directly ligate asparagine to tRNAAsn, the aspartyl‐tRNA synthetase has evolved to be specific for tRNAAsp, a discriminating aspartyl‐tRNA synthetase (D‐AspRS). How bacterial aspartyl‐tRNA synthetase evolved specificity for tRNAAsp is not clear. Bacillus subtilis encodes a ND‐AspRS while the related Bacillus anthracis codes for a D‐AspRS. The sequence similarity between the two enzymes provides an opportunity to alter the tRNA specificity of a ND‐AspRS. Using the sequence comparison along with available structures of aspartyl‐tRNA synthetases, we have mutated the anticodon binding domain of the B. subtilis ND‐AspRS and will report how the various mutations impacted specificity for tRNAAsp. The work expands our understanding of tRNA synthetase substrate specificity and how fidelity of protein synthesis is achieved.Support or Funding InformationNational Science Foundation (MCB‐1615770)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.