The binding of distinct segments of actin to multiple sites in the C-terminus of caldesmon: comparative aspects of actin interaction with troponin-I and caldesmon.

Thin-filament-based regulation of the contractile response is considered to involve the interaction of actin with troponin-I in striated muscle and the interaction of actin with caldesmon in smooth muscle. The nature of the interaction with actin of these inhibitory proteins has been studied by proton magnetic resonance spectroscopy using segments of caldesmon and troponin-I which mimic their functional properties. Caldesmon is shown to interact with two distinct sites on the N-terminal residues 1-44 of actin subdomain 1 with corresponding contacts on caldesmon domain 3 and domain 4 at its C-terminus. We demonstrate that, whereas inhibition by the troponin-I fragment (residues 96-117) is effected by its interaction with the N-terminal region of actin, the separate inhibitory ability of different regions of the C-terminus of caldesmon (domains 4a and 4b) is mediated by interaction with noncontiguous segments on subdomain 1 of actin. Our studies of the spatial relationship of these actin contacts on caldesmon further suggest that one molecule of caldesmon may associate with two actin monomers. The demonstrated interactive nature of these caldesmon attachments to distinct regions of actin is relevant to the mechanism of calcium modulation of inhibition of actomyosin ATPase by caldesmon.