Interaction of Mycobacterium tuberculosis elongation factor Tu with GTP is regulated by phosphorylation

During protein synthesis, translation elongation factor Tu (Ef-Tu) is responsible for the selection and binding of the cognate aminoacyl-tRNA to the acceptor site on the ribosome. The activity of Ef-Tu is dependent on its interaction with GTP. Post-translational modifications such as phosphorylation are known to regulate the activity of Ef-Tu in several prokaryotes. Although, a study on Mycobacterium tuberculosis phosphoproteome showed Ef-Tu to be phosphorylated, the role of phosphorylation in regulation of Ef-Tu has not been studied. In this report, we show that phosphorylation of M. tuberculosis Ef-Tu (MtbEf-Tu) by PknB reduced its interaction with GTP, suggesting a concomitant reduction in protein synthesis. Overexpression of PknB in M. smegmatis indeed reduced the protein synthesis. MtbEf-Tu was found to be phosphorylated on multiple sites by PknB including Thr118, which is required for optimal activity of the protein. We found that kirromycin, an Ef-Tu specific antibiotic, had significant effect on nucleotide binding of unphosphorylated MtbEf-Tu but not on the phosphorylated protein. Our results show that the modulation of MtbEf-Tu:GTP interaction by phosphorylation can have impact on cellular protein synthesis and growth. These results also suggest that phosphorylation can change the sensitivity of the protein to the specific inhibitors. Thus, efficacy of an inhibitor can also depend on the post-translational modification(s) of the target and should be considered during the development of the molecule