PhaR, a protein of unknown function conserved among short-chain-length polyhydroxyalkanoic acids producing bacteria, is a DNA-binding protein and represses Paracoccus denitrificans phaP expression in vitro

A putative regulatory protein, PhaR, which was identified in the polyhydrox yalkanoic acid synthetic locus (phaZCPR) in Paracoccus denitrificans, was i nvestigated. The PhaR protein purified from a recombinant Escherichia coli was estimated to be 22 kDa by sodium dodecyl sulfate-polyacrylamide gel ele ctrophoresis, being consistent with the mass calculated from the nucleotide sequence. The molecular mass was determined to be 93 kDa by size-exclusion chromatography, suggesting that the protein formed a tetramer. A gel mobil ity shirt assay showed that PhaR specifically bound to the intergenic regio n of phaC-phaP. In a cell-free protein synthesis system using E. coli S30 e xtract, the expression of the phaP gene was repressed by the addition of pu rified PhaR. These results suggest that PhaR is a DNA-binding protein and m ay play a role in the regulation of phaP gene expression (C) 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Microbiol ogical Societies.