Fig. 3. Ligand binding-induced conformation change in a protein.

a Binding of Ca²⁺ to calmodulin (CaM) causes conformation and charge changes in CaM. b Oscillation amplitude vs. U₀ plots before (red) and after (blue) Ca²⁺ binding to CaM, where the vertical bars are standard deviations of >150 CaM or Ca²⁺/CaM molecules. c Statistical analysis for 150 CaM molecules (red) and 151 Ca²⁺/CaM molecules (blue) showing the diameter (D_H), charge (q) and mobility (µ) distributions of CaM and Ca²⁺/CaM complex (see Supplementary Table 4 for a summary). The red and blue lines are Gaussian fittings of the peaks. d, f Tracking of the charge (Δq) and size (ΔD_H) changes of a single CaM molecule induced by Ca²⁺ binding over time, where the size change was measured with U₀ fixed at 7 V (the plateau regime) and the charge measurement was performed with U₀ = 4 V (the linear regime). The binding and unbinding of Ca²⁺ to CaM were performed by alternatively flowing 1 mM EGTA and 1 mM Ca²⁺ in 100× diluted PBS (at pH = 7.4) over the surface. The scatter plots (black dots) are raw data smoothed over three points, and the red lines are guide to the eye, showing the charge or size change duo the binding and unbinding of the molecule with Ca²⁺. e, g Charge and size histograms obtained from the first binding and unbinding cycle in d, f respectively. Both histograms show two peaks corresponding to the CaM molecule with and without Ca²⁺ binding.