Supporting Information

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Fig. 6. Sensorgram of WT CaM binding to skeletal muscle myosin light chain kinase (skMLCK) in the presence (dashed line) and absence (solid line) of Ca²⁺, showing that binding depends rigorously on the presence of CaM.

Fig. 7. Binding of WT CaM to skMLCK at increasing CaM concentrations. Sensorgrams are presented as the dashed lines, and the fits are presented as solid lines. Data were fit by using the BIAEVALUATION global fitting program (Biacore) according to the equation d[RL]/dt = k_a[L] – k_d[RL], yielding k_on (1.8 × 10⁵ ), k_off (3 × 10^-3), K_d (16 nM).

Fig. 8. Scatchard plot of WT (squares, solid line) and mutant (circles, dashed line) CaM binding to mutant four-charge reversal skMLCK. A linear fit to the data yielded the K_ds 270 m M (for mutant CaM) and >700 m M (for WT CaM).

Fig. 9. Stopped-flow measurement of the reaction between purified D1 protein and various concentrations of Ca²⁺. The initial reaction could be fit to a single exponential to yield the average rate (n > 3, in s^-1) for each Ca²⁺ concentration. A fit of the data (k_obs = k_on[Ca²⁺] + k_off) yields the association and dissociation rate constants.

Fig. 10. Comparison between ATP-induced Ca²⁺ oscillations in the cytosol and the endoplasmic reticulum (ER) for B cell lymphoma-2 (Bcl-2) and neo cells. It should be noted that ATP was added at time t = 0, at which point there was a large release of Ca²⁺ from the ER into the cytosol, followed by Ca²⁺ oscillations in both the ER and the cytosol. The differences between Bcl-2 (a and b) and neo (c and d) cells are highlighted by focusing on the oscillations induced by ATP. The [Ca²⁺]_cyt signal is presented in green, and the [Ca²⁺]_ER signal is presented in blue for Bcl-2 and red for neo. The conditions for the above experiments are the same as for the data presented in Fig. 4. The data are from single cells and illustrate that, although there is some variability in the oscillations in Bcl-2 and neo cells, the trends (differences in oscillation frequency and duration of release from the ER) are maintained.

Fig. 11. Structure of EC and EGCG.