title: The excluded DNA strand is SEW important for hexameric helicase unwinding.
abstract: Helicases are proposed to unwind dsDNA primarily by translocating on one strand to sterically exclude and separate the two strands. Hexameric helicases in particular have been shown to encircle one strand while physically excluding the other strand. In this article, we will detail experimental methods used to validate specific interactions with the excluded strand on the exterior surface of hexameric helicases. Both qualitative and quantitative methods are described to identify an excluded strand interaction, determine the exterior interacting residues, and measure the dynamics of binding. The implications of exterior interactions with the nontranslocating strand are discussed and include forward unwinding stabilization, regulation of the unwinding rate, and DNA damage sensing.
authors.affiliation: Molecular Biophysics and Structural Biology Program, University of Pittsburgh, Pittsburgh, PA 15260, United States.
authors.forename: Sean M
authors.initials: SM
authors.lastname: Carney
authors.affiliation: Molecular Biophysics and Structural Biology Program, University of Pittsburgh, Pittsburgh, PA 15260, United States; Department of Chemistry and Biochemistry, Baylor University Waco, TX 76798, United States. Electronic address: michael_trakselis@baylor.edu.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Trakselis
mesh_terms: D004247:DNA / Q000737:chemistry / Q000235:genetics*; D004249:DNA Damage / Q000235:genetics*; D004265:DNA Helicases / Q000737:chemistry / Q000235:genetics*; D005818:Genetic Engineering / Q000379:methods*; D009690:Nucleic Acid Conformation
chemical_list: D004247:DNA; D004265:DNA Helicases
keywords: DNA damage sensing; Helicase; Hydrogen-deuterium mass spectrometry; Single molecule FRET; Steric exclusion; Unwinding