Identification of RNA polymerase beta' subunit segment contacting the melted region of the lacUV5 promoter.
Identification of the RNA polymerase functional regions involved in interactions with promoter is a basis for understanding the mechanism of transcription initiation. We have used formaldehyde cross-linking to identify a region of Escherichia coli RNA polymerase beta' subunit contacting lacUV5 promoter in open complex. Treatment of open complex with formaldehyde results in cross-linking of beta' and sigma(70) subunits at positions -5 and -3 on the nontemplate strand of the promoter DNA. These cross-links reflect specific interactions between RNA polymerase and promoter established in open complex. The positions of formaldehyde cross-links in the beta' subunit were mapped to the N-terminal segment (Cys(198)-Met(237)), which is contiguous to the evolutionary conserved region B. The proximity of the beta' and sigma cross-links suggest that the N-terminal region of the beta' subunit, interacting with single-stranded promoter DNA, can cooperate with the sigma subunit in the process of open complex formation.
Brodolin
K
K
Institute of Molecular Genetics Russian Academy of Sciences, Kurchatov Sq. 46, Moscow 123182, Russia.
Mustaev
A
A
Severinov
K
K
Nikiforov
V
V
D000595:Amino Acid Sequence; D003432:Cross-Linking Reagents; D004269:DNA, Bacterial / Q000235:genetics* / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*
D003432:Cross-Linking Reagents; D004269:DNA, Bacterial; D012321:DNA-Directed RNA Polymerases