Identification of the Bacillus subtilis pur operon repressor

Area of Science:

• Microbiology
• Molecular Biology
• Gene Regulation

Background:

• The pur operon in Bacillus subtilis regulates purine biosynthesis.
• Transcription of this operon is repressed when adenine is abundant.

Purpose of the Study:

• To identify and characterize the regulatory protein and gene responsible for pur operon repression in Bacillus subtilis.
• To elucidate the mechanism of adenine-mediated repression.

Main Methods:

• Purification of the repressor protein.
• Cloning and overexpression of the purR gene.
• In vitro protein-DNA binding assays.
• In vivo transcription analysis using mutant strains.

Main Results:

• Purified the PurR repressor, a 62-kDa homodimer that binds to the pur operon control region.
• Identified the PurR binding site (~110 bp) overlapping the promoter.
• Demonstrated that 5-phosphoribosyl 1-pyrophosphate inhibits PurR binding.
• Showed that mutations in the binding site or purR abolish repression in vivo and binding in vitro.
• Found that purR is autoregulated.

Conclusions:

• Established a model where excess adenine signals PurR through the 5-phosphoribosyl 1-pyrophosphate pool to regulate pur operon transcription.
• Highlighted the lack of similarity between Bacillus subtilis and Escherichia coli purine repressors.