Conserved region 2.1 of Escherichia coli heat shock transcription factor σ32 is required for modulating both metabolic stability and transcriptional activity
Conserved region 2.1 of Escherichia coli heat shock transcription factor σ32 is required for modulating both metabolic stability and transcriptional activity
复制标题
DOI：
10.1128/jb.186.22.7474-7480.2004
复制
发表时间：
2004-11-01
影响因子：
3.2
通讯作者：
Kanemori, M
中科院分区：
文献类型：
--
作者：
Horikoshi, M;Yura, T;Kanemori, M
大肠杆菌热休克转录因子西格玛(32)在体内降解迅速，半衰期约为1分钟。一组蛋白质，包括DNAK伴侣组(DNAK、DNAJ、GRPE)和ATP依赖的蛋白酶(FtsH、HsIUV等)。参与了西格玛(32)的降解。为了进一步了解sigma(32)稳定性的调节，我们分离了明显稳定的sigma(32)突变体。许多突变体在2.1区域的N端半部分(残基47到55)有氨基酸替换，这是一个在细菌或因子中高度保守的区域。半衰期比野生型蛋白质长约2倍到10倍以上。除了更稳定之外，在产生稳定西格玛的细胞中，热休克蛋白水平，如DNAK和GroEL，也增加了(32)。详细分析表明，一些稳定的sigma(32)突变体具有比野生型更高的转录活性。这些结果表明，2.1区域的N端半部分是调节代谢稳定性和Sigma(32)活性所必需的。证据表明，sigma(32)的稳定并不是由于对核心RNA聚合酶亲和力的提高。因此，区域2.1可能参与与蛋白质降解机制的相互作用，包括分子伴侣。
Escherichia coli heat shock transcription factor sigma(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, HsIUV, etc.) are involved in degradation of sigma(32). To gain further insight into the regulation of sigma(32) stability, we isolated sigma(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 or 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 sigma(32). Detailed analysis showed that some stable sigma(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 sigma(32). The evidence suggests that sigma(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.