Atomic structure of and valine binding to the regulatory ACT domain of the Mycobacterium tuberculosis Rel protein. (3rd November 2020)

• Record Type: Journal Article
• Authors: Shin, Joon; Singal, Bharti; Sony Subramanian Manimekalai, Malathy; Wei Chen, Ming; Ragunathan, Priya; Grüber, Gerhard
• Is Part Of: FEBS journal. Volume 288:Number 7(2021)
• Page Start: 2377
• Page End: 2397
• Publication Date: 2020-11-03
• Subjects: ACT domain -- Mycobacterium tuberculosis -- NMR spectroscopy -- Rel -- stringent response -- tuberculosis; Biochemistry -- Periodicals; Molecular biology -- Periodicals; Pathology, Molecular -- Periodicals; 572
• DOI: 10.1111/febs.15600
• Languages: English
• ISSNs: 1742-464X

Abstract

Abstract : The stringent response, regulated by the bifunctional (p)ppGpp synthetase/hydrolase Rel in mycobacteria, is critical for long‐term survival of the drug‐tolerant dormant state of Mycobacterium tuberculosis . During amino acid starvation, Mt Rel senses a drop in amino acid concentration and synthesizes the messengers pppGpp and ppGpp, collectively called (p)ppGpp. Here, we investigate the role of the regulatory 'Aspartokinase, Chorismate mutase and TyrA' (ACT) domain in Mt Rel. Using NMR spectroscopy approaches, we report the high‐resolution structure of dimeric Mt Rel ACT which selectively binds to valine out of all other branched‐chain amino acids tested. A set of Mt Rel ACT mutants were generated to identify the residues required for maintaining the head‐to‐tail dimer. Through NMR titrations, we determined the crucial residues for binding of valine and show structural rearrangement of the Mt Rel ACT dimer in the presence of valine. This study suggests the direct involvement of amino acids in (p)ppGpp accumulation mediated by Mt Rel independent to interactions with stalled ribosomes. Database Structural data are available in the PDB database under the accession number 6LXG . Abstract : The stringent response (SR) is crucial for survival as well as optimal growth of Mycobacterium tuberculosis (Mtb) . Mycobacterial cells initiate SR during nutrient and energy limitations by increasing levels of (p)ppGpp. In Mtb, (p)ppGpp synthesis and degradation are carried out by the bifunctional enzyme Rel. Here, we present the dimeric solution structure of its ACT domain, unraveled its specific binding to valine, and determined the critical residues in Val‐ACT binding.