Roles of N10 formyl tetrahydrofolate N10 fTHF ribosomal large subunit pseudouridine synthase D RluD and transcription translation coupling in the fidelity of translation initiation in Escherichia coli

Abstract

Protein synthesis is a fundamental process in gene expression. It involves four basic steps initiation, elongation, termination, and ribosome recycling in bacteria. However, the step of initiation is the most regulated and rate-limiting step. Regulation at this step can save cellular energy and avoid toxicity due to the accretion of defective proteins. Bacteria have evolved with several mechanisms to ensure the fidelity of translation initiation. One of these mechanisms is the selection of the initiator tRNA (i-tRNA) at the P-site of the ribosome. The i-tRNA is endowed with unique structural features which facilitate its selection at the P-site. Two of the special properties of the i-tRNAs are the formylation of amino acid attached to it and, the highly conserved occurrence of the three consecutive G-C (3GC) base pairs in its anticodon stem. While the property of formylation facilitates its binding to the 30S ribosome, the 3GC pairs are important for its recognition in the ribosomal P-site and the maintenance of the fidelity of translation initiation. However, the 3GC base pairs mediated mechanisms used by the cell and the ribosome to ensure the fidelity of initiation are not fully understood. The mutations in the 3GC base pairs avert its selection at the P-site and its involvement in initiation. We made use of this phenotype in isolating E. coli mutants which allow translation initiation with the mutant 3GC i-tRNA. The characterization of many of the mutation(s) in these isolates has revealed the factors or pathways used by the cell to ensure the selection of canonical i-tRNA for initiation. Previously, the characterization of some of these suppressors unveiled that not only the ribosomal or the translation factors but also that cellular metabolism plays a vital role in ensuring the fidelity of translation initiation...