A β-hairpin is a Minimal Latch that Supports Positive Supercoiling by Reverse Gyrase

Abstract

Reverse gyrase is a unique type I topoisomerase that catalyzes the introduction of positive supercoils into DNA in an ATP-dependent reaction. Supercoiling is the result of a functional cooperation of the N-terminal helicase domain with the C-terminal topoisomerase domain. The helicase domain is a nucleotide-dependent conformational switch that alternates between open and closed states with different affinities for single- and double-stranded DNA. The isolated helicase domain as well as full-length reverse gyrase can transiently unwind double-stranded regions in an ATP-dependent reaction. The latch region of reverse gyrase, an insertion into the helicase domain with little conservation in sequence and length, has been proposed to coordinate events in the helicase domain with strand passage by the topoisomerase domain. Latch deletions lead to a reduction in or complete loss of supercoiling activity. Here we show that the latch consists of two functional parts, a globular domain that is dispensable for DNA supercoiling and a β-hairpin that connects the globular domain to the helicase domain and is required for supercoiling activity. The β-hairpin thus constitutes a minimal latch that couples ATP-dependent processes in the helicase domain to DNA processing by the topoisomerase domain.