Identification of the promoter region for the colanic acid polysaccharide biosynthetic genes in Escherichia coli K-12

Abstract

The colanic acid polysaccharide capsule biosynthetic genes (cps genes) are primarily clustered at one site located at about 45 min on the Escherichia coli chromosome. The network of proteins involved in regulating the expression of these genes includes the two positive regulators RcsA and RcsB. This work describes the site of action of these two activator proteins and the promoter of the cps genes. It is likely that the cps genes are arranged in a single long operon that is at least 13.5 kb. The promoter region was identified with fusions to lacZ that resulted in regulated expression by the Rcs network of regulatory proteins, and the start site of transcription was identified by primer extension. The operator region was cloned from Kohara phage to multicopy plasmids and identified by titrating RcsA or RcsB. Sequence analysis of the promoter and operator region revealed homology to the JUMPstart element found in the untranslated region of many exopolysaccharide biosynthetic operons. In addition, the deduced amino acid sequence of the amino terminus of the first open reading frame of the cps operon was found to be homologous to proteins encoded by the exopolysaccharide biosynthetic operons of Klebsiella pneumoniae and Erwinia amylovora.