Properties of the Escherichia coli DNA-binding (unwinding) protein interaction with nucleolytic enzymes and DNA

Abstract

The Escherichia coli DNA-binding protein reduces the activity of most single-stranded DNases from a variety of sources; however, the nucleases associated with E. coli DNA polymerase II and bacteriophage T7-induced DNA polymerase and E. coli exonuclease I remain active in the presence of binding protein and may even be stimulated. Those nucleases whose activities are stimulated by binding protein interact with the protein in the absence of DNA, to give a specific enzyme-binding protein complex, e.g. E. coli DNA polymerase II and E. coli exonuclease I both form a species, the sedimentation of which is compatible with a complex of one enzyme molecule plus one or two monomers of binding protein. Furthermore, the apparent affinity of nucleolytic enzymes for their substrate, single-strand DNA, is altered by complexing the DNA with binding protein. In particular, DNA polymerase II binds in a stable way only to DNA that is saturated with binding protein. This facilitation of binding to DNA led to the observation that in contrast to the mechanism of digestion of free DNA, E. coli DNA polymerase II digests DNA complexed with DNA-binding protein that is in a manner in keeping with a processive mechanism.