ENERGY COUPLING IN ESCHERICHIA-COLI DNA GYRASE - THE RELATIONSHIP BETWEEN NUCLEOTIDE-BINDING, STRAND PASSAGE, AND DNA SUPERCOILING

Binding of the nonhydrolyzable ATP analogue 5'-adenylyl-beta,gamma-imi dodiphosphate (ADPNP) to Escherichia coli DNA gyrase can lead to a lim ited noncatalytic supercoiling of DNA, Here we examine the efficiency of coupling between ADPNP binding and the change in linking number eit her of positively or negatively supercoiled plasmid DNA or of small DN A circles, The coupling efficiency varies from 100% (Delta Lk = -2 per gyrase tetramer, a stoichiometry of I) with positively supercoiled su bstrates under certain reaction conditions to an undetectably low valu e with moderately negatively supercoiled substrates (sigma = -0.046) o r small circular substrates. Furthermore, the rate of ADPNP binding to the gyrase-DNA complex is also dependent on the topological state of the DNA; the previously observed slow binding of ADPNP to the complex of gyrase with linear DNA is accelerated 16-fold when the substrate DN A is negatively supercoiled, suggesting a functional interaction betwe en the nucleotide-binding and DNA-binding domains which is independent of the strand-passage process. The implications for the normal ATP-de pendent supercoiling reaction of the enzyme are considered and the res ults discussed in terms of current mechanistic models for DNA gyrase a ction and the possible in vivo roles of the enzyme.