Binding and activation of DNA topoisomerase III by the Rmi1 subunit

Abstract

Rmi1 is a conserved oligonucleotide and oligosaccharide binding-fold protein that is associated with RecQ DNA helicase complexes from humans (BLM-TOP3α) and yeast (Sgs1-Top3). Although human RMI1 stimulates the dissolution activity of BLM-TOP3α, its biochemical function is unknown. Here we examined the role of Rmi1 in the yeast complex. Consistent with the similarity of top3Δ and rmi1Δ phenotypes, we find that a stable Top3·Rmi1 complex can be isolated from yeast cells overexpressing these two subunits. Compared with Top3 alone, this complex displays increased superhelical relaxation activity. The isolated Rmi1 subunit also stimulates Top3 activity in reconstitution experiments. In both cases elevated temperatures are required for optimal relaxation unless the substrate contains a single-strand DNA (ssDNA) bubble. Interestingly, Rmi1 binds only weakly to ssDNA on its own, but it stimulates the ssDNA binding activity of Top3 5-fold. Top3 and Rmi1 also cooperate to bind the Sgs1 N terminus and promote its interaction with ssDNA. These results demonstrate that Top3-Rmi1 functions as a complex and suggest that Rmi1 stimulates Top3 by promoting its interaction with ssDNA. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc.