title: Intramolecular synapsis of duplex DNA by vaccinia topoisomerase.
abstract: Complexes formed by vaccinia topoisomerase I on plasmid DNA were visualized by electron microscopy. The enzyme formed intramolecular loop structures in which non-contiguous DNA segments were synapsed within filamentous protein stems. At high enzyme concentrations the DNA appeared to be zipped up within the protein filaments such that the duplex was folded back on itself. Formation of loops and filaments was also observed with an active site mutant, Topo-Phe274. Binding of Topo-Phe274 to relaxed DNA circles in solution introduced torsional strain, which, after relaxation by catalytic amounts of wild-type topo-isomerase, resulted in acquisition of negative supercoils. We surmise that the topoisomerase-DNA complex is a plectonemic supercoil in which the two duplexes encompassed by the protein filaments are interwound in a right handed helix. We suggest that topoisomerase-mediated DNA synapsis plays a role in viral recombination and in packaging of the 200 kbp vaccinia genome during virus assembly.
authors.affiliation: Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10021, USA.
authors.forename: S
authors.initials: S
authors.lastname: Shuman
authors.forename: D G
authors.initials: DG
authors.lastname: Bear
authors.forename: J
authors.initials: J
authors.lastname: Sekiguchi
mesh_terms: D001665:Binding Sites; D004247:DNA / Q000378:metabolism* / Q000648:ultrastructure; D004264:DNA Topoisomerases, Type I / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D004270:DNA, Circular / Q000378:metabolism / Q000648:ultrastructure; D004278:DNA, Superhelical / Q000378:metabolism / Q000648:ultrastructure; D008854:Microscopy, Electron; D017354:Point Mutation; D014616:Vaccinia virus / Q000201:enzymology* / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D004270:DNA, Circular; D004278:DNA, Superhelical; D014764:Viral Proteins; D004247:DNA; D004264:DNA Topoisomerases, Type I