Estimating the parameters of a four-site binding model. (A) CaM-binding data from multiple experimental groups (adapted from Stefan et al., 2009). M_i refers to CaM with i-bound calcium ions. (B) Parameters obtained by fitting the sequential binding model in A to experimental data from five groups. Parameters are from Linse et al., 1991 (squares), Haiech et al., 1981 (rhombi), Porumb, 1994 (circles), Burger et al., 1984 (triangles), and Crouch and Klee, 1980 (inverted triangles). (C) Synthetic, noiseless binding data (solid curve) calculated using parameters from Linse et al. (1991). (D) Two distinct parameter sets that yield excellent fits to the data. For parameters shown as squares, all binding sites have nearly identical affinity, consistent with weak cooperativity. The parameters shown as circles are consistent with strong cooperative interactions between the binding sites. The binding curves generated from these parameters are plotted in C as circles and squares.