M Zylicz

The dnaK protein modulates the heat-shock response of Escherichia coli

E. coli bacteria respond to a sudden upward shift in temperature by transiently overproducing a small subset of their proteins, one of which is the product of the dnaK gene. Mutations in dnaK have been previously shown to affect both DNA and RNA synthesis in E. coli. Bacteria carrying the dnaK756 mutation fail to turn off the heat-shock response at 43 degrees C. Instead, they continue to synthesize the heat-shock proteins in large amounts and underproduce other proteins. Both reversion and P1 transduction analyses have shown that the failure to turn off the heat-shock response is the result of the dnaK756 mutation. In addition, bacteria that overproduce the dnaK protein at all temperatures undergo a drastically reduced heat-shock response at high temperature. We conclude that the dnaK protein is an inhibitor of the heat-shock response in E. coli.

Bacteriophage λ replication proteins: Formation of a mixed oligomer and binding to the origin of λ DNA

SummaryThe purified bacteriophage λ replication proteins O and P sediment separately in metrizamide gradients of low ionic strength as dimers. Together they interact with each other forming an oligomer, composed of two molecules of λO and one molecule of λP. The λO-P oligomer is active in the in vitro replication of oriλ-containing DNA.Equilibrium sedimentation in preformed metrizamide density gradients under conditions that separate DNA-protein complexes from free proteins was employed in order to study possible interactions among the λ replication proteins and oriλ DNA. It was found that the λP protein binds specifically to oriλ-containing plasmid DNA only in the presence of λO protein. About 100 molecules of λO and 10 molecules of λP form a complex with the oriλ DNA. The λ DNA-λO-λP complex was shown to be active in an in vitro replication system.Since the physical interactions between oriλ and λO and between λP and the Escherichia coli dnaB replication protein are well documented, the evidence for a λO-P interaction presented in this paper provides the missing link in the molecular mechanism that enables λ to direct the host replication machinery to the replication of its own DNA.