Oral Presentation 16th Asian Conference on Transcription 2019

Structure - Function Relationship of omega- subunit of RNA polymerase: Through the lens of silent mutations  (977)

Dipankar Chatterji 1
  1. Indian Institute of Science, Bangalore, KARNATAKA, India

RNA polymerase  is composed of a total of 6 subunits. Information is available on the contribution of all the subunits in the primary functioning of bacterial RNA polymerase except the smallest subunit omega. A toxic mutant screen was used to discover the role of omega in RNAP holoenzyme. This led to the serendipitous discovery of a silent mutant which was lethal. The primary focus of our study is to decode the mechanism behind this lethality. The native omega is intrinsically disordered but the silent mutant was having a predominantly helical structure. So, we hypothesized that this transition from intrinsically disordered to ordered structure is responsible for the lethal phenotype and the disordered nature of omega is prerequisite for maintaining the plasticity of the active site.  We generated several silent mutants of omega to investigate the role of codon bias and effect of rare codons with respect to its position in the protein sequence. Not all silent mutations affect the structure equally. RNA polymerase reconstituted with these omega silent mutants (structured in comparison to wild type) were found to be inactive. Concomitant changes in structure and transcriptional profile led to understand that proteins with same sequence can have different fold and function. Codon plus strain was used to see the rescue of phenotype in lethal silent mutants.  We performed ribosome profiling experiments with the wild type and mutant strains of ω and observed that they are translated at a differential rate. ppGpp interaction studies also showed difference.