PMID:7783622
| Citation |
Quail, MA and Guest, JR (1995) Purification, characterization and mode of action of PdhR, the transcriptional repressor of the pdhR-aceEF-lpd operon of Escherichia coli. Mol. Microbiol. 15:519-29 |
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| Abstract |
The repressor of the pdhR-aceEF-lpd operon of Escherichia coli, PdhR, was amplified to 23% of total cell protein and purified to homogeneity by heparin-agarose and cation-exchange chromatography. The purified protein is a monomer (M(r) 29,300) which binds specifically to DNA fragments containing the pdh promoter (Ppdh) in the absence of pyruvate. The pdh operator was identified by DNase I footprinting as a region of hyphenated dyad symmetry, +11AATTGGTaagACCAATT+27, situated just downstream of the transcript start site. In vitro transcription from Ppdh was repressed > 1000-fold by PdhR and this repression was antagonized in a concentration-dependent manner by its co-effector, pyruvate. Studies on RNA polymerase binding at Ppdh showed that RNA polymerase protects the -44 to +21 region in the absence of PdhR, but no RNA polymerase binding or protection upstream of +9 could be detected in the presence of PdhR. It is concluded that PdhR represses transcription by binding to an operator site centred at +19 such that effective binding of RNA polymerase is prevented. |
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| Keywords |
Acetyltransferases/biosynthesis; Acetyltransferases/genetics; Bacterial Proteins/biosynthesis; Bacterial Proteins/genetics; Bacterial Proteins/isolation & purification; Base Sequence; DNA, Bacterial/genetics; DNA, Bacterial/metabolism; DNA-Directed RNA Polymerases/metabolism; Dihydrolipoamide Dehydrogenase/biosynthesis; Dihydrolipoamide Dehydrogenase/genetics; Dihydrolipoyllysine-Residue Acetyltransferase; Escherichia coli/genetics; Escherichia coli/metabolism; Escherichia coli Proteins; Gene Expression Regulation, Bacterial/drug effects; Ketoglutarate Dehydrogenase Complex/biosynthesis; Ketoglutarate Dehydrogenase Complex/genetics; Molecular Sequence Data; Operon/genetics; Pyruvate Dehydrogenase Complex/biosynthesis; Pyruvate Dehydrogenase Complex/genetics; Pyruvates; Pyruvic Acid; Recombinant Fusion Proteins/metabolism; Repressor Proteins/genetics; Repressor Proteins/isolation & purification; Transcription Factors/genetics; Transcription Factors/isolation & purification |
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