Multiple regions on the Escherichia coli heat shock transcription factor sigma32 determine core RNA polymerase binding specificity.
We have analyzed the core RNA polymerase (RNAP) binding activity of the purified products of nine defective alleles of the rpoH gene, which encodes sigma32 in Escherichia coli. All mutations studied here lie outside of the putative core RNAP binding regions 2.1 and 2.2. Based on the estimated K(s)s for the mutant sigma and core RNAP interaction determined by in vitro transcription and by glycerol gradient sedimentation, we have divided the mutants into three classes. The class III mutants showed greatly decreased affinity for core RNAP, whereas the class II mutants' effect on core RNAP interaction was only clearly seen in the presence of sigma70 competitor. The class I mutant behaved nearly identically to the wild type in core RNAP binding. Two point mutations in class III altered residues that were distant from one another. One was found in conserved region 4.2, and the other was in a region conserved only among heat shock sigma factors. These data suggest that there is more than one core RNAP binding region in sigma32 and that differences in contact sites probably exist among sigma factors.
Joo
D M
DM
Department of Molecular and Cell Biology, University of California, Berkeley 94720, USA.
Nolte
A
A
Calendar
R
R
Zhou
Y N
YN
Jin
D J
DJ
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D017354:Point Mutation; D012808:Sigma Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D006360:Heat-Shock Proteins; D012808:Sigma Factor; D014157:Transcription Factors; C069774:heat-shock sigma factor 32; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases