Computational analysis of the effect of a binding protein (RbpA) on the dynamics of Mycobacterium tuberculosis RNA polymerase assembly

RNA polymerase-binding protein A (RbpA) is an actinomycetes-specific protein crucial for the growth and survival of the pathogen Mycobacterium tuberculosis. Its role is essential and influences the transcription and antibiotic responses. However, the regulatory mechanisms underlying RbpA-mediated transcription remain unknown. In this study, we employed various computational techniques to investigate the role of RbpA in the formation and dynamics of the RNA polymerase complex. Our analysis reveals significant structural rearrangements in RNA polymerase happen upon interaction with RbpA. Hotspot residues, crucial amino acids in the RbpA-mediated transcriptional regulation, were identified through our examination. The study elucidates the dynamic behavior within the complex, providing insights into the flexibility and functional dynamics of the RbpA-RNA polymerase interaction. Notably, potential allosteric mechanisms, involving the interface of subunits 1 and 2 were uncovered, shedding light on how RbpA modulates transcriptional activity. Finally, potential ligands meant for the 1-2 binding site were identified through virtual screening. The outcomes of our computational study serve as a foundation for experimental investigations into inhibitors targeting the RbpA-regulated dynamics in RNA polymerase. Overall, this research contributes valuable information for understanding the intricate regulatory networks of RbpA in the context of transcription and suggests potential avenues for the development of RbpA-targeted therapeutics. Author SummaryInfection studies by Mycobacterium tuberculosis (Mtb) acquires primary importance due to its severe infection and antibiotic resistance. There is an open need for highly effective drugs and one needs to employ novel approaches such as detailed structural analysis and the possibility to focus on allosteric inhibitors. We have exploited the availability of cryo-EM structures of RNA polymerase of Mtb, with and without its transcription-activator protein namely RNA polymerase-binding protein A (RbpA). In this study, we employed various computational techniques to investigate the role of RbpA in the formation and dynamics of the RNA polymerase complex. The assemblies were subject to molecular dynamics and perturbation scanning, followed by structural comparisons and measurement of subunit interface strength. These analyses could clearly show that subunits, which are far away from the RbpA binding site, undergo differential structural changes. Hence, we focused on the site to recognize potential small molecule inhibitors using virtual screening. These analyses demonstrate that it is possible to perform comparative structural analysis of different forms of assemblies, which can be useful towards drug design.