Figure 5. Probing the final contour length increment with Poly (GS) inserts.
(A) Structural homology model overlay of wild-type and mutant Doc containing a (GS)8-linker between helix 1 and helix 3. The wild-type Doc is shown in green. The 16 amino acid long GS-insert is shown in purple (Kelley and Sternberg, 2009) (remaining Doc domain not shown). (B) Typical force extension trace with final double rupture event depicted in green (arrow). (C) Histogram and kernel density estimate of the transformation of the single force extension trace in panel B into contour length space (black) and kernel density estimate of the whole dataset of single molecule Xyn-Doc:Coh-CBM traces bearing xylanase fingerprint and final double rupture (gray, offset in y-direction for readability) in contour length space. (D) Histograms (bars, bin width: 1 nm), kernel density estimates (drawn lines, bandwidth: 0.75 nm, gaussian kernel), and statistical test (Kolmogorov-Smirnov, ‘KS test’) are each calculated on the raw data of the final increments (peak-to-peak distances) in contour length space (x-distance between arrow 1 and 2 in panel (C). Maxima for final double event increments lie at 7.75 nm and 7.73 nm for iLOV-Coh:Doc (wild-type)-sfGFP (N = 255) and Xyn-Doc (GS)8:Coh-CBM (N = 320) final ruptures, respectively (a two-sample KS test on the raw data indicates no significant difference in the data distributions (p-value of 21.7%).