Structure function relationship in the omega subunit of escherichia coli RNA polymerase

Abstract

Gene expression in all cells involves synthesis of RNA from a DNA template newlinewhich is carried out by a multi-subunit enzyme known as DNA-dependent RNA newlinepolymerase. In the eubacteria, this enzyme is made up of four subunits, a, p, pt, and newlinero, which comprise the core polymerase. newlineThe smallest subunit ro consists of only 90 amino acids and has a molecular newlineweight of 10,105 Da. Although a subunit of RNAP, its function was still elusive. newlineUnlike the other subunits the ro subunit is non-essential in E. coli. However, omega newlinehomologs are present in most bacteria whose genomes have been sequenced and are newlinealso present in archaebacteria and eukaryotes such as yeast, drosophila and even newlinehumans. Sequence analysis shows the presence of three regions of homology, namely, newlineCR1, CR2 and CR3 suggesting some important conserved function. newlineThe first indication of the function of ro was the demonstration that ro-less newlineRNAP was transcriptionally less active as compared to the wild type polymerase. newlineRenaturation of ro-less RNAP in the presence of ro led to a restoration in its activity to newlinethat of the wild type. Moreover the ro-less RNAP was associated with large amounts newlineof GroEL. This led to the speculation that ro might have a GroEL like chaperone newlinefunction in maintaining the structure of RNA polymerase. newlineHowever the exact function or the structure of ro was still not elucidated. newlineHence this study was taken up to investigate the structure and function of the newlinedispensable subunit, ro, in greater detail. newlineChapter 1 introduces the structure-function relationship among different newlinesubunits of RNA polymerase as well as the concept of molecular chaperones. newlineiii newlineThe structural organization of the ro subunit was explored in Chapter 2. For newlinethis purpose we have used the technique of limited proteolysis. Thjs has revealed the newlinepresence of a 53 amino acid domain in theN-terminal portion of ro that has all the newlinestructure and function of the full-length protein. newlineThe previous work prompted the assignment of a chaperone like function for newlinero.