CRISPR–Cas9 Mediated DNA Unwinding Detected Using Site-Directed Spin Labeling

The RNA-guided CRISPR–Cas9 nuclease has revolutionized genome engineering, yet its mechanism for DNA target selection is not fully understood. A crucial step in Cas9 target recognition involves unwinding of the DNA duplex to form a three-stranded R-loop structure. Work reported here demonstrates direct detection of Cas9-mediated DNA unwinding by a combination of site-directed spin labeling and molecular dynamics simulations. The results support...