Fine balance in the regulation of DnaB helicase by DnaC protein in replication in Escherichia coli

Abstract

The DnaC protein of Escherichia coli is essential for replication in vivo and in vitro. In the initiation of replication of a minichromosome at its origin, DnaC delivers the DnaB helicase from a DnaB.DnaC complex to the future replication fork and then departs. However, if an excess of DnaC was present in subsequent steps, it severely inhibited replication by slowing the DnaB helicase at the replication fork. When DnaB was present at a level equimolar with the excess DnaC, the inhibition was relieved, implying that the ratio of DnaC to DnaB is critical for achieving optimal replication activity and avoiding inhibition by DnaC. In vivo, overproduction of DnaC slowed cell growth. This slowing was alleviated by overproducing DnaB at the same time. E. coli strains with a dnaCts gene defective in chromosomal initiation were complemented by the wild-type gene in trans. On the other hand, strains with an elongation-defective dnaCts gene were not complemented by the wild-type dnaC gene. The dominance of the mutant protein suggests that it remains tightly complexed with DnaB at the replication fork, inhibiting elongation even in the presence of the wild-type DnaC.