title: DNA sequence alignment by microhomology sampling during homologous recombination.
abstract: Homologous recombination (HR) mediates the exchange of genetic information between sister or homologous chromatids. During HR, members of the RecA/Rad51 family of recombinases must somehow search through vast quantities of DNA sequence to align and pair single-strand DNA (ssDNA) with a homologous double-strand DNA (dsDNA) template. Here, we use single-molecule imaging to visualize Rad51 as it aligns and pairs homologous DNA sequences in real time. We show that Rad51 uses a length-based recognition mechanism while interrogating dsDNA, enabling robust kinetic selection of 8-nucleotide (nt) tracts of microhomology, which kinetically confines the search to sites with a high probability of being a homologous target. Successful pairing with a ninth nucleotide coincides with an additional reduction in binding free energy, and subsequent strand exchange occurs in precise 3-nt steps, reflecting the base triplet organization of the presynaptic complex. These findings provide crucial new insights into the physical and evolutionary underpinnings of DNA recombination.
authors: Department of Biochemistry and Molecular Biophysics, Columbia University, 650 West 168(th) Street, New York, NY 10032, USA.
authors: Zhi
authors: Z
authors: Qi
authors: Department of Chemistry, Columbia University, 650 West 168(th) Street, New York, NY 10032, USA.
authors: Sy
authors: S
authors: Redding
authors: Department of Biochemistry and Molecular Biophysics, Columbia University, 650 West 168(th) Street, New York, NY 10032, USA.
authors: Ja Yil
authors: JY
authors: Lee
authors: Department of Biochemistry and Molecular Biophysics, Columbia University, 650 West 168(th) Street, New York, NY 10032, USA.
authors: Bryan
authors: B
authors: Gibb
authors: Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
authors: YoungHo
authors: Y
authors: Kwon
authors: Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
authors: Hengyao
authors: H
authors: Niu
authors: Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
authors: William A
authors: WA
authors: Gaines
authors: Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
authors: Patrick
authors: P
authors: Sung
authors: Department of Biochemistry and Molecular Biophysics, Columbia University, 650 West 168(th) Street, New York, NY 10032, USA; Howard Hughes Medical Institute, Columbia University, 650 West 168(th) Street, New York, NY 10032, USA. Electronic address: ecg2108@columbia.edu.
authors: Eric C
authors: EC
authors: Greene
mesh_terms: D018797:Cell Cycle Proteins / Q000378:metabolism; D023902:Chromosome Pairing; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D059765:Homologous Recombination; D051135:Rad51 Recombinase / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: D018797:Cell Cycle Proteins; C513033:DMC1 protein, S cerevisiae; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; C495151:RAD51 protein, S cerevisiae; D051135:Rad51 Recombinase