Figure 2. Effect of the central helix mutations on the Ca²⁺ binding properties of isolated cTnC.

Panel A shows increases in Trp fluorescence, which occur as Ca²⁺ binds to the N-domains of cTnC (■), cTnC^D87A/D88A (●), or cTnC^{E94A/E95A/E96A} (○) at 15°C. The cTnC proteins contained F27W substitution. The Trp fluorescence was excited at 285 nm and monitored at 345 nm. Panels B and C show the time course of decreases in Trp fluorescence as Ca²⁺ was removed by excess EGTA from the N-domains of isolated cTnC, cTnC^D87A/D88A, or cTnC^{E94A/E95A/E96A}. Panel B: measured at 15°C. Panel C: measured at 5°C. The data traces have been normalized and staggered for clarity. Each trace is an average of at least five traces fit with a single exponential equation. The Trp fluorescence was excited at 285 nm and monitored through a WG320 filter.