The Escherichia coli C homoprotocatechuate degradative operon: hpc gene order, direction of transcription and control of expression

Summary

Homoprotocatechuate (HPC; 3,4-dihydroxyphenylacetate) is catabolized to Krebs cycle intermediates via extradiol (meta-) cleavage and the necessary enzymes are chromosomally encoded in a variety of bacteria. Based on an analysis of the cloned pathway genes, the Escherichia coli C hpc gene cluster was thought to be arranged in two gene blocks transcribed from a central, divergent, operator/promoter region, which was negatively regulated by the Hpc repressor. By a variety of techniques including expression of cloned hpc genes in pUC18/19 vectors, unidirectional deletion subcloning, hybridization studies and nucleotide sequencing it has now been shown that the hpc pathway structural genes are transcribed in one direction. These experiments have also indicated that a decarboxylase and an isomerase of the pathway are encoded by a single gene (hpcE) and have established the exact structural gene order as hpcRphpcECBDGH. The position of the putative regulatory gene, hpcR, is upstream of the first structural gene (hpcE) for the Hpc pathway enzymes. The deduced open reading frame for the Hpc repressor specifies a protein of 148 amino acids with a subunit molecular weight of 17 kDa. The region between hpcR and the first gene for the pathway enzymes has a sequence similar to that for catabolite activator protein (CAP) binding. This region is immediately upstream of a promoter for the pathway structural genes, which has been identified by transcript mapping.