Conserved Region 2.1 of Escherichia coli Heat Shock Transcription Factor σ 32 Is Required for Modulating both Metabolic Stability and Transcriptional Activity

Abstract

ABSTRACT Escherichia coli heat shock transcription factor σ 32 is rapidly degraded in vivo, with a half-life of about 1 min. A set of proteins that includes the DnaK chaperone team (DnaK, DnaJ, GrpE) and ATP-dependent proteases (FtsH, HslUV, etc.) are involved in degradation of σ 32 . To gain further insight into the regulation of σ 32 stability, we isolated σ 32 mutants that were markedly stabilized. Many of the mutants had amino acid substitutions in the N-terminal half (residues 47 to 55) of region 2.1, a region highly conserved among bacterial σ factors. The half-lives ranged from about 2-fold to more than 10-fold longer than that of the wild-type protein. Besides greater stability, the levels of heat shock proteins, such as DnaK and GroEL, increased in cells producing stable σ 32 . Detailed analysis showed that some stable σ 32 mutants have higher transcriptional activity than the wild type. These results indicate that the N-terminal half of region 2.1 is required for modulating both metabolic stability and the activity of σ 32 . The evidence suggests that σ 32 stabilization does not result from an elevated affinity for core RNA polymerase. Region 2.1 may, therefore, be involved in interactions with the proteolytic machinery, including molecular chaperones.