Heme binding to calmodulin, troponin C, and parvalbumin, as a probe of calcium-dependent conformational changes.

Heme-CO binds to the active (calcium-bound) form of calmodulin (CaM), but not to the inactive form. Despite a similarity in structure of another calcium-binding protein, skeletal muscle troponin C, both the affinity and the spectral red-shift of the absorption of the heme group are greatly decreased for troponin C relative to calmodulin. Parvalbumin, another calcium-binding protein, shows a twofold greater affinity for heme-CO relative to CaM. Unlike calmodulin and troponin C, the affinity of parvalbumin for heme-CO is even greater in the absence of calcium. The affinity of the tryptic and thrombic fragments of CaM for heme-CO are decreased relative to the entire calmodulin. The binding of heme-CO is specific as demonstrated by the discrimination of the calmodulin, troponin C, and parvalbumin pockets. The interaction of heme-CO with active (calcium-bound) CaM is rapid (ms) as determined by stopped flow measurements. No difference in kinetics was observed for mixing inactive (calcium free) CaM with a solution of [heme-CO plus calcium], indicating that the calcium-binding step and subsequent change in protein conformation are rapid.