DNA transport by a type II DNA topoisomerase: evidence in favor of a two-gate mechanism

Abstract

DNA substrates in which a supercoiled DNA is singly linked to a nicked or relaxed DNA ring were used to analyze the transport of one DNA ring through another by yeast DNA topoisomerase II. The enzyme binds preferentially to the supercoiled DNA and promotes decatenation efficiently upon binding of a nonhydrolyzable ATP analog. Analysis of the reaction products shows that the nicked or relaxed DNA ring released is not associated with the enzyme-supercoiled DNA complex. These results favor a two-gate model in which the DNA ring being transported can exit from the interior of the enzyme through a gate on the opposite side of the entrance gate, which is irreversibly closed upon binding of the nonhydrolyzable ATP analog.