Nucleotide sequence of korB, a replication control gene of broad host-range plasmid RK2.

The korB gene is a major regulatory element in the replication and maintenance of broad host-range plasmid RK2. It negatively controls the replication gene trfA, the host-lethal determinants kilA and kilB, and the korA-korB operon. Here, we present the nucleotide sequence of an 1167 base-pair region that encodes korB. Using sequence data from korB mutants, we identified the korB structural gene. The predicted polypeptide product is negatively charged and has a molecular weight of 39,015, which is considerably less than that estimated by its electrophoretic mobility in SDS/polyacrylamide gels. Secondary-structure predictions of korB polypeptide revealed three closely spaced helix-turn-helix regions with significant homology to similar structures in known DNA-binding proteins. The korB gene, like all other sequenced RK2 genes, shows a strong preference for codons ending in a G or C residue. This is similar to codon usage by genes of Klebsiella and Pseudomonas, the original hosts for RK2 and some closely related plasmids. We also sequenced the site of transposon Tn76 insertion in the host-range mutant pRP761 and found it to be located immediately upstream from korB in the incC gene. Finally, we report the presence of sequences resembling a replication origin within the korB structural gene: a cluster of four 19 base-pair direct repeats and a nearby potential binding site for Escherichia coli dna A replication protein.