A repressor protein, PhaR, regulates polyhydroxyalkanoate (PHA) synthesis via its direct interaction with PHA.

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• Phasins (PhaP) are proteins associated with polyhydroxyalkanoate (PHA) granules, enhancing PHA synthesis.
• The phaR gene, located downstream of phaP in Paracoccus denitrificans, encodes a negative regulator of PhaP expression.

Purpose of the Study:

• To investigate the regulatory role of PhaR in PhaP expression and PHA synthesis.
• To elucidate the mechanism by which PhaR senses PHA granules.

Main Methods:

• DNase I footprinting to identify PhaR binding sites upstream of phaP and phaR.
• Recombinant Escherichia coli expressing PhaR to study its interaction with PHA granules.
• In vitro expression experiments to assess PhaR-mediated repression and P(3HB) derepression of phaP.

Main Results:

• PhaR binds specifically to two regions upstream of phaP and phaR, indicating roles in both phaP regulation and autoregulation.
• PhaR binds directly to poly[(R)-3-hydroxybutyrate] [P(3HB)] granules, and P(3HB) or oligomers cause PhaR dissociation from DNA.
• PhaP expression is repressed by PhaR and derepressed by P(3HB), suggesting PhaR senses PHA synthesis onset and granule enlargement.

Conclusions:

• PhaR acts as a sensor for PHA synthesis and granule development through direct binding to PHA.
• A PhaR-mediated regulatory model for PHA synthesis is proposed, involving sensing of PHA granules.
• The widespread distribution of PhaR homologs suggests a conserved and crucial role in regulating PHA synthesis in bacteria.