2024/11/29 by Gillian M. Cadden, Jan-Gero Schloetel, Grant McKenzie +3 · 1 voice
Biochemistry, Genetics and Molecular Biology · Chemistry · #Advanced biosensing and bioanalysis techniques #Biochemistry #Biology #Biophysics #Chemistry #Crystallography #DNA #DNA Repair Mechanisms #DNA and Nucleic Acid Chemistry #Enzyme #Fluorescence #Förster resonance energy transfer #Genetics #Physics #Protein subunit #Recombinase #Recombination #Tetramer
paper · pdf · doi:10.1038/s41467-024-54531-4
openalex publication_date 2024/11/29 · openalex created_date 2025/10/10 · openalex updated_date 2026/07/23
Abstract Serine recombinases are proposed to catalyse site-specific recombination by a unique mechanism called subunit rotation. Cutting and rejoining DNA occurs within an intermediate synaptic complex comprising a recombinase tetramer bound to two DNA sites. After double-strand cleavage at both sites, one half of the complex rotates 180° relative to the other, before re-ligation of the DNA ends. We used single-molecule FRET (smFRET) methods to provide compelling direct physical evidence for subunit rotation by recombinases Tn3 resolvase and Sin. Synaptic complexes containing fluorescently labelled DNA show FRET fluctuations consistent with the subunit rotation model. FRET changes were associated with the rotation steps, on a timescale of 0.4–1.1 s-1 <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML"> <mml:msup> <mml:mrow> <mml:mstyle> <mml:mtext>s</mml:mtext> </mml:mstyle> </mml:mrow> <mml:mrow> <mml:mo>−</mml:mo> <mml:mn>1</mml:mn> </mml:mrow> </mml:msup> </mml:math> , as well as opening and closing of the gap between the scissile phosphates during cleavage and ligation. Multiple rounds of recombination were observed within the ~25 s observation period, including frequent consecutive rotation events in the cleaved-DNA state without evidence of intermediate ligation.