Fig. 5
From: Single molecule high-throughput footprinting of small and large DNA ligands
Mechanical footprint (MFP) for restriction enzymes. a Schematics of the binding of a large protein, such a restriction enzyme, on a DNA sequence. In the general case of asymmetric binding, depending on the enzyme orientation and on which part of the enzyme binds to the recognition sequence, the shift between the measured blockage position during hairpin unzipping (corresponding to the head or back of the enzyme) and the recognition site will be different. The MFP, corresponding to the DNA size covered by the enzyme, is computed as the sum of the shifts measured in the two enzyme orientations plus the size of the restriction site. In the particular case of symmetric binding, the central part of the enzyme binds to the recognition site, and the measured shifts from the head and back of the enzyme are expected to be the same. The MFP is then computed as twice the measured shift plus the size of the restriction site. b Schematics of recognition and cleavage of the palindromic Rsa1 restriction enzyme, as well as its binding to hairpins H s4. c Distribution of blockage positions obtained by applying the FC protocol to the H s4 hairpin in presence of the Rsa1 enzyme (number of beads = 72). Error bars are inversely proportional to the square root of the number of points for each bin. A single blockage is observed that is shifted 5 bps away from the recognition site. The MFP is estimated 14 bp, twice the shift plus the 4 bps corresponding to the restriction site
