Assignment of the positions of chymotryptic fragments and cysteinyl groups in the primary structure of caldesmon in relation to a conformational change.

Caldesmon is universally associated with smooth muscle thin filaments, and reportedly interacts with actin, calmodulin, tropomyosin, and myosin. I attempted to determine the positions of the chymotryptic fragments which have been used to study the sites of such interactions in its primary structure. Such assignment, combined with the accumulated data of fragment studies, made it possible to clarify the functional domain organization of the caldesmon molecule. Using a specific cleavage method involving nitrothiocyanobenzoate, I also determined the number and locations of cysteinyl residues in the amino-acid sequence. Two cysteinyl groups thus determined were located close to the ends and almost at the same distance apart from the N- and C-termini of the whole molecule, respectively. Taking these locations and the extraordinary sensitivity to oxidation into consideration, I could reasonably elucidate the origin of the controversial data and the interpretation so far reported from various laboratories, concerning the length and conformation of the native molecule. Caldesmon might be folded in solution and its contour length could be almost double the accepted value, which was hydrodynamically estimated.