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J Bacteriol, March 1998, p. 1095-1102, Vol. 180, No. 5
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Multiple Regions on the Escherichia coli Heat Shock Transcription Factor sigma 32 Determine Core RNA Polymerase Binding Specificity

Daniel M. Joo,1,dagger Audrey Nolte,1 Richard Calendar,1,* Yan Ning Zhou,2 and Ding Jun Jin2

Department of Molecular and Cell Biology, University of California, Berkeley, California 94720,1 and Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 208922

Received 29 September 1997/Accepted 23 December 1997

We have analyzed the core RNA polymerase (RNAP) binding activity of the purified products of nine defective alleles of the rpoH gene, which encodes sigma 32 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 Kss 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 sigma 70 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 sigma 32 and that differences in contact sites probably exist among sigma factors.

* Corresponding author. Mailing address: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720-3202. Phone: (510) 642-5951. Fax: (510) 643-5035. E-mail: rishard{at}socrates.berkeley.edu.

dagger Present address: Department of Microbiology and Immunology, University of California, San Francisco, CA 94143.




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