Fig. 2.

AFM unfolding of hN2-NRR region. (A) Raw data from AFM experiments with NRR showing distinct unfolding features corresponding to the LNR (Left) and HD domains (Right). Worm-like chain (WLC) analysis shown on unfolding curves (black line). Mean coefficient of determination for WLC analysis of data = 0.98. The first peak was shown (via controls) to correspond to salt contacts between the tip and slide, and is therefore not analyzed by the WLC. Frequency of extension (B) and force (C) observed during all unfolding events (LNR and HD) in the presence (striped; n = 1,075) and absence (block color; n = 866) of coordinated ions (x axis values represent upper limit of histogram interval). Curve on extension histogram highlights the peaks for LNR domains unfolding. Distinct HD domain unfolding not observed (expected extension 55 nm). Variations due to nonspecific attachments to the hN2-NRR construct outside the polyK tag produce background features over a large range of extensions. Curves on the force histogram highlight bimodalality and therefore unfolding of two species. In the absence of ions, this bimodal feature is lost and the force required for unfolding is reduced.