Dual Pathways for B. anthracis Asparaginyl‐tRNA Formation

Abstract

In Bacillus anthracis, there are two distinct routes for attaching asparagine (Asn) to its cognate transfer RNA (tRNAAsn), an essential step in protein synthesis. In the direct route, asparaginyl‐tRNA synthetase (AsnRS) directly ligates Asn to tRNAAsn. The indirect pathway is a two‐step process where a non‐discriminating‐AspRS (ND‐AspRS) attaches aspartate (Asp) to tRNAAsn which is them amidated by GatCAB, an amidotransferase. In this pathway, Asn is ultimately synthesized on the tRNA using a complex called the transamidosome which is made up of the ND‐AspRS, tRNAAsn, and GatCAB. B. anthracis encodes two distinct AspRSs with one of bacterial origins and the other acquired through horizontal gene transfer from archaea. The archaeal AspRS serves as the non‐discriminating enzyme for tRNA‐dependent asparagine biosynthesis. We have successfully overproduced in E. coli and purified the enzymes for both pathways to further characterize the two routes under various physiologically relevant conditions. The research provides a better understanding as to why this pathogen acquired an archaeal aspartyl‐tRNA synthetase and encodes two distinct pathways for asparaginyl‐tRNA synthesis.