CALCIUM-DEPENDENT REGULATION OF THE CALDESMON HEAVY-MEROMYOSIN INTERACTION BY CALTROPIN

The binding of chicken gizzard caldesmon to smooth muscle heavy meromy osin (HMM) was studied using caldesmon-Sepharose 4B affinity chromatog raphy, far-ultraviolet circular dichroism (CD), and the fluorescent pr obe acrylodan. When HMM was applied to a caldesmon-Sepharose column in the presence of 40 mM NaCl, most of the protein was retained on the c olumn, and HMM could be eluted by increasing the NaCl level to 0.5 M; this interaction was not Ca2+-dependent. Far-UV CD studies indicated a n interaction between caldesmon and HMM since the experimentally obser ved ellipticity values at 222 and 207 nm deviated from the theoretical values for the complex, and this interaction was also not Ca2+-sensit ive. Addition of HMM to a caldesmon-caltropin complex induced a confor mational change suggesting the formation of a ternary complex for whic h Ca2+ was essential. Acrylodan-labeled caldesmon, when excited at 375 nm, had an emission maximum at 515 +/- 2 nm. Addition of HMM resulted in a nearly 20% decrease in fluorescence intensity with little or no shift in the emission maximum. Titration of HMM with labeled caldesmon indicated a strong affinity for HMM [K(a) was on the order of (4.5 +/ - 0.5) x 10(7) M-1], and this interaction was observed both in the pre sence and in the absence of calcium. When HMM was titrated with labele d caldesmon in the presence of caltropin in a 0.2 mM Ca2+ medium, its affinity for caldesmon was lowered nearly 3-fold [K(a) almost-equal-to (1.50 +/- 0.5) x 10(7) M-1]. Caltropin, which is very potent in rever sing the inhibitory effect of caldesmon in the presence of calcium (Ma ni et al., 1992), is shown in this study to modulate the interaction b etween caldesmon and smooth muscle heavy meromyosin, thus making it a potential calcium factor in regulating caldesmon in smooth muscle.