title: Different quaternary structures of human RECQ1 are associated with its dual enzymatic activity.
abstract: RecQ helicases are essential for the maintenance of chromosome stability. In addition to DNA unwinding, some RecQ enzymes have an intrinsic DNA strand annealing activity. The function of this dual enzymatic activity and the mechanism that regulates it is, however, unknown. Here, we describe two quaternary forms of the human RECQ1 helicase, higher-order oligomers consistent with pentamers or hexamers, and smaller oligomers consistent with monomers or dimers. Size exclusion chromatography and transmission electron microscopy show that the equilibrium between the two assembly states is affected by single-stranded DNA (ssDNA) and ATP binding, where ATP or ATPgammaS favors the smaller oligomeric form. Our three-dimensional electron microscopy reconstructions of human RECQ1 reveal a complex cage-like structure of approximately 120 A x 130 A with a central pore. This oligomeric structure is stabilized under conditions in which RECQ1 is proficient in strand annealing. In contrast, competition experiments with the ATPase-deficient K119R and E220Q mutants indicate that RECQ1 monomers, or tight binding dimers, are required for DNA unwinding. Collectively, our findings suggest that higher-order oligomers are associated with DNA strand annealing, and lower-order oligomers with DNA unwinding.
authors.affiliation: International Centre for Genetic Engineering and Biotechnology, Trieste, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Muzzolini
authors.forename: Fabienne
authors.initials: F
authors.lastname: Beuron
authors.forename: Ardan
authors.initials: A
authors.lastname: Patwardhan
authors.forename: Venkateswarlu
authors.initials: V
authors.lastname: Popuri
authors.forename: Sheng
authors.initials: S
authors.lastname: Cui
authors.forename: Benedetta
authors.initials: B
authors.lastname: Niccolini
authors.forename: Mathieu
authors.initials: M
authors.lastname: Rappas
authors.forename: Paul S
authors.initials: PS
authors.lastname: Freemont
authors.forename: Alessandro
authors.initials: A
authors.lastname: Vindigni
mesh_terms: D001667:Binding, Competitive; D002850:Chromatography, Gel; D004247:DNA / Q000378:metabolism*; D006801:Humans; D008854:Microscopy, Electron; D020836:Protein Structure, Quaternary; D053484:RecQ Helicases / Q000378:metabolism*
chemical_list: D004247:DNA; D053484:RecQ Helicases