| dc.description.abstract | Proteins interact with other proteins to maintain functional homeostasis of the cell by tightly regulating cellular processes. Hence, it becomes important to understand not only the downstream effects of protein-protein interactions (PPIs), but its impact on interacting partners too. Much of the work embodied in this thesis pertains to sequence, structure, dynamics and functional analysis of protein-protein complexes. Firstly, the impact of PPIs on structure and dynamics of individual interacting partners was studied using a dataset of 58 protein-protein complexes of known 3-D structure. It was observed that binding elicits change in structure and dynamics of many interacting proteins, not only at the interface, but also at the regions away from the interfaces. Analyses of various examples showed that such changes could a) contribute towards the stability of the complex by adding to the positive gain in entropy, b) regulate a downstream function such as binding of another protein and c) alter the functional capacity of an enzyme. Next, this learning was extended to a protein-protein complex between DNA gyrase and CcdB toxin from E. coli to understand the dissociation of the former due to binding of another protein, the CcdA antitoxin. It was observed that binding of CcdA to the CcdB-gyrase complex elicits a series of response in the CcdB toxin. This response could be recorded in terms of change in residue flexibility/conformation at strategic locations, that facilitates release of gyrase from the assembly. The toxin-antitoxin (TA) systems are small, genetic elements composed of a toxin gene (encoding toxin protein) and a neighbouring antitoxin gene (encoding antitoxin protein) regulated under the same operon. These specific protein pairs from M. tuberculosis (Mtb) were analysed to understand the basis of their unique expansion. Sequence analysis of VapBC and MazEF TA systems of Mtb suggested similarities in their binding modes and functional residues. Structure of known TA complexes were utilised to detect interface as well as substrate-binding residues, which were further validated by laboratory experiments performed by the collaborators. To check the uniqueness of Mtb TA systems, their homologues from other micro-organisms were identified. Interestingly, while most orthologues were found well-conserved in the members of the Mtb complex, the soil-inhabiting, free-living Actinobacteria also harboured as many as 12 toxin-antitoxin pairs. A detailed analysis of Mtb TA systems helped in identifying sequence features of these systems. Further, these features along with TA features from other organisms led to the identification of four novel TA systems, namely, Rv2515c-Rv2514c, Rv3642c-Rv3641c, Rv0367c-Rv0366c, and Rv0023-Rv0024. Detailed computational analysis supported by laboratory experiments (performed by collaborators) suggested novel VapBC system for Rv2515c-Rv2514c and non-canonical PezAT for Rv0367c-Rv0366c. Lastly, because there exist many TA systems in Mtb, attempts were made to explore the potential of these TA systems to cross-react with one another. To this end, extensive analyses of experimental and modelled structures and promoter sequences of TA systems were performed. The results obtained helped in identification of specificity conferring residues in VapB15 and MazEFs and enabled proposition of cross-talks. Taken together, results presented in this thesis shed light on the characteristics of specific PPIs and how they evolve, function and impact their partners to control cellular processes. | en_US |
