FIGURE 9.

sCaM1 and sCaM4 response to slow Ca²⁺ transients. (A) Time course of change in intrinsic Trp fluorescence in the NADK peptide as sCaM1 is transiently occupied by Ca²⁺ in the presence of Mg²⁺. 3 μM WT sCaM1 with 9 μM NADK peptide, 1 mM Mg²⁺ and 500 μM EDTA in stopped-flow buffer (10 mM MOPS, 150 mM KCl, pH 7.0) plus 1mM Mg²⁺ was rapidly mixed with an equal volume of increasing Ca²⁺ [0 μM (black), 10 μM (red), 30 μM (green), 60 μM (cyan), 120 μM (blue), 250 μM (magenta), 2 mM (orange)] in stopped-flow buffer plus 1 mM Mg²⁺ at 20°C. (B) Time course of change in intrinsic Trp fluorescence in NADK as sCaM4 is transiently occupied by Ca²⁺ in the presence of Mg²⁺. 3 μM WT sCaM4 with 9 μM NADK, 1 mM Mg²⁺ and 500 μM EDTA in stopped-flow buffer plus 1 mM Mg²⁺ was rapidly mixed with an equal volume of increasing Ca²⁺ (0, 10, 30, 60, 120, and 250 μM, 2 mM) in stopped-flow buffer plus 1 mM Mg²⁺ at 20°C. There was no significant difference between sCaM1 and sCaM4 occupancy under these conditions. The visible occupancy for all traces was determined as described in section “Materials and Methods.”