PMID:9878422
| Citation |
Jovanovic, G, Rakonjac, J and Model, P (1999) In vivo and in vitro activities of the Escherichia coli sigma54 transcription activator, PspF, and its DNA-binding mutant, PspFDeltaHTH. J. Mol. Biol. 285:469-83 |
|---|---|
| Abstract |
Transcription of the phage-shock protein (psp) operon in Escherichia coli is driven by a sigma54 promoter, stimulated by integration host factor and dependent on an upstream, cis-acting sequence and an activator protein, PspF. PspF belongs to the enhancer binding protein family but lacks an N-terminal regulatory domain. Purified PspF is not modified and has an ATPase activity that is increased twofold in the presence of DNA carrying the psp cis-acting sequence. Purified mutant His-tagged PspF that lacks the C-terminal DNA-binding motif has a DNA-independent ATPase activity when present at 30-fold the concentration of the wild-type protein. Both proteins oligomerize in solution in an ATP and DNA-independent manner. The wild-type activator protein, but not the DNA-binding mutant, binds specifically to the cis-acting sequence. Analysis of the sequence protected by PspF demonstrates the presence of two upstream binding sites within the sequence, UAS I and UAS II, which together constitute the psp enhancer. Protection at low protein concentrations is more pronounced and more extensive on a supercoiled DNA than on a linear template. Full expression of the psp operon upon hyperosmotic shock depends on wild-type PspF, but only partially requires the presence of the psp enhancer. |
| Links |
PubMed Online version:10.1006/jmbi.1998.2263 |
| Keywords |
Adenosine Triphosphatases/metabolism; Bacterial Proteins/genetics; Bacterial Proteins/isolation & purification; Bacterial Proteins/metabolism; Base Sequence; Binding Sites; DNA, Bacterial; DNA-Binding Proteins/genetics; DNA-Binding Proteins/isolation & purification; DNA-Binding Proteins/metabolism; DNA-Directed RNA Polymerases/metabolism; Enhancer Elements, Genetic; Escherichia coli/genetics; Escherichia coli Proteins; Heat-Shock Proteins/genetics; Histidine; Molecular Sequence Data; Mutagenesis; Operon; Osmotic Pressure; Promoter Regions, Genetic; RNA Polymerase Sigma 54; Recombinant Fusion Proteins/genetics; Recombinant Fusion Proteins/isolation & purification; Recombinant Fusion Proteins/metabolism; Sigma Factor/metabolism; Trans-Activators/genetics; Trans-Activators/isolation & purification; Trans-Activators/metabolism |
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