title: DNA G-segment bending is not the sole determinant of topology simplification by type II DNA topoisomerases.
abstract: DNA topoisomerases control the topology of DNA. Type II topoisomerases exhibit topology simplification, whereby products of their reactions are simplified beyond that expected based on thermodynamic equilibrium. The molecular basis for this process is unknown, although DNA bending has been implicated. To investigate the role of bending in topology simplification, the DNA bend angles of four enzymes of different types (IIA and IIB) were measured using atomic force microscopy (AFM). The enzymes tested were Escherichia coli topo IV and yeast topo II (type IIA enzymes that exhibit topology simplification), and Methanosarcina mazei topo VI and Sulfolobus shibatae topo VI (type IIB enzymes, which do not). Bend angles were measured using the manual tangent method from topographical AFM images taken with a novel amplitude-modulated imaging mode: small amplitude small set-point (SASS), which optimises resolution for a given AFM tip size and minimises tip convolution with the sample. This gave improved accuracy and reliability and revealed that all 4 topoisomerases bend DNA by a similar amount: ~120° between the DNA entering and exiting the enzyme complex. These data indicate that DNA bending alone is insufficient to explain topology simplification and that the 'exit gate' may be an important determinant of this process.
authors.affiliation: Department of Oral Biology, School of Dentistry and Molecular and Nanoscale Physics Group, School of Physics and Astronomy, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Neil H
authors.initials: NH
authors.lastname: Thomson
authors.affiliation: 1] Department of Oral Biology, School of Dentistry and Molecular and Nanoscale Physics Group, School of Physics and Astronomy, University of Leeds, Leeds, LS2 9JT, United Kingdom [2].
authors.forename: Sergio
authors.initials: S
authors.lastname: Santos
authors.affiliation: Department of Biological Chemistry, John Innes Centre Norwich Research Park, Norwich NR4 7UH, United Kingdom.
authors.forename: Lesley A
authors.initials: LA
authors.lastname: Mitchenall
authors.affiliation: 1] Department of Biological Chemistry, John Innes Centre Norwich Research Park, Norwich NR4 7UH, United Kingdom [2].
authors.forename: Tanya
authors.initials: T
authors.lastname: Stuchinskaya
authors.affiliation: 1] Department of Biological Chemistry, John Innes Centre Norwich Research Park, Norwich NR4 7UH, United Kingdom [2].
authors.forename: James A
authors.initials: JA
authors.lastname: Taylor
authors.affiliation: Department of Biological Chemistry, John Innes Centre Norwich Research Park, Norwich NR4 7UH, United Kingdom.
authors.forename: Anthony
authors.initials: A
authors.lastname: Maxwell
mesh_terms: D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D008970:Molecular Weight; D009690:Nucleic Acid Conformation; D011485:Protein Binding
chemical_list: D004247:DNA; D004250:DNA Topoisomerases, Type II