Enhanced DNA Scanning and Target Specificity via Motif Sequestration

DNA-binding proteins require effective targeting of specific sequences amidst complex genomes without engaging in unproductive interactions with non-target regions. Researchers have discovered that the site-specific recombinase Cre experiences an autoinhibitory, conformational state which diminishes its affinity for non-specific DNA, facilitating linear scanning while suppressing the formation of recombinogenic complexes.

Studies have indicated that the C-terminal portion of Cre adopts a temporary, secluded conformation that blocks the catalytic domain's binding to DNA. Through the use of NMR spectroscopy, single-molecule fluorescence microscopy, and optical trapping, researchers examined how this conformational equilibrium influences target search.

NMR measurements indicate that the sequestered state is more abundant in the absence of DNA and remains at non-specific sites, while interaction with the correct loxP sequences releases the autoinhibition. Single-molecule tracking reveals that the entire Cre protein undergoes continuous one-dimensional diffusion and becomes firmly bound to its target sites, while truncation of the C-terminal segment hampers scanning efficiency by halting diffusion.

These findings reveal how temporary motif sequestration synchronizes target search with the assembly of site-specific complexes, suggesting a general mechanism by which DNA-modifying enzymes strike a balance between search efficiency and target specificity.