Specific recognition of the -10 promoter element by the free RNA polymerase sigma subunit.
Bacterial RNA polymerase holoenzyme relies on its sigma subunit for promoter recognition and opening. In the holoenzyme, regions 2 and 4 of the sigma subunit are positioned at an optimal distance to allow specific recognition of the -10 and -35 promoter elements, respectively. In free sigma, the promoter binding regions are positioned closer to each other and are masked for interactions with the promoter, with sigma region 1 playing a role in the masking. To analyze the DNA-binding properties of the free sigma, we selected single-stranded DNA aptamers that are specific to primary sigma subunits from several bacterial species, including Escherichia coli and Thermus aquaticus. The aptamers share a consensus motif, TGTAGAAT, that is similar to the extended -10 promoter. We demonstrate that recognition of this motif by sigma region 2 occurs without major structural rearrangements of sigma observed upon the holoenzyme formation and is not inhibited by sigma regions 1 and 4. Thus, the complex process of the -10 element recognition by RNA polymerase holoenzyme can be reduced to a simple system consisting of an isolated sigma subunit and a short aptamer oligonucleotide.
Sevostyanova
Anastasiya
A
Institute of Molecular Genetics, Russian Academy of Sciences, Kurchatov Sq, 2, Moscow, Russia.
Feklistov
Andrey
A
Barinova
Nataliya
N
Heyduk
Ewa
E
Bass
Irina
I
Klimasauskas
Saulius
S
Heyduk
Tomasz
T
Kulbachinskiy
Andrey
A
D001426:Bacterial Proteins / Q000378:metabolism; D016384:Consensus Sequence; D003432:Cross-Linking Reagents; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism*; D034301:Deinococcus / Q000201:enzymology; D004926:Escherichia coli / Q000201:enzymology; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D021122:Protein Subunits / Q000378:metabolism; D013824:Thermus / Q000201:enzymology
D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D021122:Protein Subunits; D012321:DNA-Directed RNA Polymerases