Binding and activation of DNA topoisomerase III by the Rmi1 subunit.
Rmi1 is a conserved oligonucleotide and oligosaccharide binding-fold protein that is associated with RecQ DNA helicase complexes from humans (BLM-TOP3 alpha) and yeast (Sgs1-Top3). Although human RMI1 stimulates the dissolution activity of BLM-TOP3 alpha, its biochemical function is unknown. Here we examined the role of Rmi1 in the yeast complex. Consistent with the similarity of top3Delta and rmi1Delta 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.
Chen
Chi-Fu
CF
Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New Jersey 08854.
Brill
Steven J
SJ
Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New Jersey 08854. Electronic address: brill@mbcl.rutgers.edu.
D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D004264:DNA Topoisomerases, Type I / Q000235:genetics / Q000378:metabolism*; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D006801:Humans; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D053484:RecQ Helicases / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D002352:Carrier Proteins; D004271:DNA, Fungal; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; C501527:RMI1 protein, human; C502206:Rmi1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C502374:TOP3 protein, S cerevisiae; C090472:SGS1 protein, S cerevisiae; D053484:RecQ Helicases; D004264:DNA Topoisomerases, Type I
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