A cycle of binding and release of the DnaK, DnaJ and GrpE chaperones regulates activity of the Escherichia coli heat shock transcription factor sigma32.

Abstract

The chaperone system formed by DnaK, DnaJ and GrpE mediates stress‐dependent negative modulation of the Escherichia coli heat shock response, probably through association with the heat shock promoter‐specific sigma32 subunit of RNA polymerase. Interactions of the DnaK system with sigma32 were analysed. DnaJ and DnaK bind free, but not RNA polymerase‐bound, sigma32 with dissociation constants of 20 nM and 5 muM respectively. Association and dissociation rates of DnaJ‐sigma32 complexes are 5900‐ and 20‐fold higher respectively than those of DnaK‐sigma32 complexes in the absence of ATP. ATP destabilizes DnaK‐sigma32 interactions. DnaJ, through rapid association with sigma32 and stimulation of hydrolysis of DnaK‐bound ATP, mediates efficient binding of DnaK to sigma32 in the presence of ATP, resulting in DnaK‐DnaJ‐sigma32 complexes containing ADP. GrpE binding to these complexes stimulates nucleotide release and subsequent complex dissociation by ATP. We propose that the principles of this cycle also operate in other chaperone activities of the DnaK system. DnaK and DnaJ cooperatively inhibit sigma32 activity in heat shock gene transcription and GrpE partially reverses this inhibition. These data indicate that reversible inhibition of sigma32 activity through transient association of DnaK and DnaJ is a central regulatory element of the heat shock response.

The hrcA and hspR regulons of Campylobacter jejuni

“…Escherichia coli and many other proteobacteria use the alternative sigma factor s32 (Grossman et al, 1984;Yura & Nakahigashi, 1999), which directs RNA polymerase to promoters of heat-shock genes, and is regulated by a number of mechanisms including a feedback loop that reduces the HSR once the stress has ended (Gamer et al, 1996;Tomoyasu et al, 1998;El-Samad et al, 2005). Most bacteria use one or more of a number of repressors to regulate the HSR (Narberhaus, 1999).…”

A 70-kDa molecular chaperone, DnaK, from the industrial bacterium Bacillus licheniformis: gene cloning, purification and molecular characterization of the recombinant protein

“…To date, the majority of works that have been done on defi ned DnaK-DnaJ systems at a prokaryotic level has principally involved in the E. coli system [18][19][20][21][22]. A number of works have also been performed on other prokaryotic organisms [23][24][25][26][27][28][29].…”

Region 2.1 of the Escherichia coli heat-shock sigma factor RpoH (σ 32) is necessary but not sufficient for degradation by the FtsH protease

“…Some proteins might share overlapping binding sites because interaction of some partners is mutually exclusive. Incubation of RpoH with DnaK or GroEL recruits the sigma factor away from the RNAP and inhibits transcription initiation in vitro (Gamer et al, 1996;Guisbert et al, 2004;Liberek et al, 1992). Conversely, in complex with RNAP the sigma factor is protected from attack by the FtsH protease in vitro (Blaszczak et al, 1999;Urech et al, 2000).…”

“…DnaK, DnaJ, GrpE and GroESL have been shown to interact with RpoH (Gamer et al, 1996;Guisbert et al, 2004;Liberek et al, 1992). One possible explanation for the stabilization of RpoH mutated in region 2.1 might be an impaired interaction with the chaperones.…”

“…The activity and stability of RpoH is modulated by the chaperone systems DnaKJ and GroESL (Gamer et al, 1996;Guisbert et al, 2004;Straus et al, 1990). At physiological temperature, e.g.…”