Abstract
Ribonucleotide analogs bound in the initiating site of Escherichia coli RNA polymerase-promoter complex were cross-linked to the beta subunit. Using limited proteolysis and chemical degradation, the cross-link was mapped to a segment of beta between amino acids Val516 and Arg540. This region (Rif-cluster I) is known to harbor many rifampicin-resistant (RifR) mutations. The results demonstrate that Rif-culster I is part of the "5'-face" of the active center and provide structural basis for the long-known effects of RifR mutations on transcription initiation, elongation, and termination.
Highlights
The cellular multisubunit DNA-dependent RNA polymerase is the central enzyme of gene expression and a target for genetic regulation
The best studied RNA polymerase (RNAP) is that of Escherichia coli
In order to identify amino acid residues participating in the active center of RNAP, we took the approach of mapping the sites of chemical cross-linking of substrate analogs
Summary
The cellular multisubunit DNA-dependent RNA polymerase is the central enzyme of gene expression and a target for genetic regulation. The assumption rested on the Rif ability to block extension of short initial transcripts [8], the failure of RifR mutants to bind Rif [9], and elongation defects observed in certain RifR mutants (10 –12). All of these effects could be explained by allosteric mechanisms whereby RifR mutations and/or Rif would exert their effects at a distance. There is still no rigorous proof that mutationally defined Rif-clusters in the middle of  participate directly in Rif binding site or in the RNAP active center. Our present results directly position Rif-cluster I at the 5Ј face of the initiating NTP site
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
