Different Quaternary Structures of Human RECQ1 Are Associated with Its Dual Enzymatic Activity

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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 ATPγS favors the smaller oligomeric form. Our three-dimensional electron microscopy reconstructions of human RECQ1 reveal a complex cage-like structure of approximately 120 Å × 130 Å 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. The transient opening of the DNA double helix is a fundamental step in several DNA metabolic processes. This reaction is driven by proteins called helicases, which make use of ATP as fuel to unwind the DNA duplex. The RecQ family of helicases help maintain genome stability. Recent studies have shown that RecQ helicases, in addition to promoting DNA unwinding, can also catalyze the opposite reaction—the pairing of the partially unwound DNA duplexes. The mechanisms underlying the regulation of this dual enzymatic activity are, however, unknown. Here we describe two structural forms of the human RECQ1 helicase, a large oligomeric complex composed of five or six subunits, and a smaller form consistent with only one or two molecules. We provide an initial view of the three-dimensional structure of the larger complex and show that this state is associated with DNA strand annealing, whereas the smaller form carries out DNA unwinding. The functional switch from strand-annealing to DNA unwinding is controlled by ATP binding, which promotes the dissociation of the larger, higher-order complexes. By providing insight into the mechanisms regulating RecQ helicase activity, our study opens a new window onto a fundamental aspect of DNA metabolism.