Regulation of the isophthalate catabolic operon controlled by IphR in Comamonas sp. strain E6

Abstract

The isophthalate (IPA) catabolic operon (iphACBDR) of Comamonas sp. strain E6 responsible for the conversion of IPA into protocatechuate is negatively regulated by an IclR-type transcriptional regulator, IphR. Promoter analysis showed that the region sufficient for the IPA-dependent induction of the iphA promoter was located within the 87 bp region upstream from the iphA start codon. The transcription start site of the iph operon was mapped at a cytosine located 49 bp upstream of the iphA start codon. Two inverted repeat sequences IR1 (positions -21 to -7 relative to the iphA transcription start site) and IR2 (-2 to +10) were found in the binding region of IphR identified by electrophoretic mobility shift assays (EMSA) using purified IphR. Mutation analysis of each half-site of these inverted repeat sequences indicated that both IR2 and a downstream half-site of IR1 are necessary for the binding of IphR. EMSA in the presence of IPA or its analogous substrates demonstrated that IPA had the ability to inhibit the binding of IphR to this operator region. In conclusion, the iph operon is negatively autoregulated by the binding of IphR to the operator region, and this repression is released by the presence of IPA.