RecQ helicase stimulates both DNA catenation and changes in DNA topology by topoisomerase III.
Together, RecQ helicase and topoisomerase III (Topo III) of Escherichia coli comprise a potent DNA strand passage activity that can catenate covalently closed DNA (Harmon, F. G., DiGate, R. J., and Kowalczykowski, S. C. (1999) Mol. Cell 3, 611-620). Here we directly assessed the structure of the catenated DNA species formed by RecQ helicase and Topo III using atomic force microscopy. The images show complex catenated DNA species involving crossovers between multiple double-stranded DNA molecules that are consistent with full catenanes. E. coli single-stranded DNA-binding protein significantly stimulated both the topoisomerase activity of Topo III alone and the DNA strand passage activity of RecQ helicase and Topo III. Titration data suggest that an intermediate of the RecQ helicase unwinding process, perhaps a RecQ helicase-DNA fork, is the target for Topo III action. Catenated DNA is the predominant product under conditions of molecular crowding; however, we also discovered that RecQ helicase and single-stranded DNA-binding protein greatly stimulated the intramolecular strand passage ("supercoiling") activity of Topo III, as revealed by changes in the linking number of uncatenated DNA. Together our results demonstrate that RecQ helicase and Topo III function together to comprise a potent and concerted single-strand DNA passage activity that can mediate both catenation-decatenation processes and changes in DNA topology.
Harmon
Frank G
FG
Division of Biological Sciences, Section of Microbiology, Center for Genetics and Development, University of California-Davis, 1 Shields Avenue, Davis, CA 95616, USA.
Brockman
Joel P
JP
Kowalczykowski
Stephen C
SC
D000251:Adenosine Triphosphatases / Q000502:physiology*; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004265:DNA Helicases / Q000502:physiology*; D004264:DNA Topoisomerases, Type I / Q000502:physiology*; D043864:DNA, Catenated / Q000096:biosynthesis* / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000502:physiology; D029968:Escherichia coli Proteins / Q000502:physiology; D007700:Kinetics; D018625:Microscopy, Atomic Force; D009690:Nucleic Acid Conformation; D011092:Polyethylene Glycols / Q000494:pharmacology; D053484:RecQ Helicases; D017186:Titrimetry
D043864:DNA, Catenated; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D011092:Polyethylene Glycols; D004247:DNA; D000251:Adenosine Triphosphatases; C506724:RecQ protein, E coli; D004265:DNA Helicases; D053484:RecQ Helicases; D004264:DNA Topoisomerases, Type I