ALIGNMENT OF CALDESMON ON THE ACTIN-TROPOMYOSIN FILAMENTS

We have reported previously that each smooth-muscle caldesmon binds pr edominantly to a region within residues 142-227 of tropomyosin, but a weaker binding site also exists at the N-terminal region of tropomyosi n [Watson, Kuhn, Novy, Lin and Mak (1990) J. Biol. Chem. 265, 18860-18 866]. In view of recent evidence for the presence of tropomyosin-bindi ng sites at both the N- and C-terminal domains of caldesmon, we have s tudied the binding of the N- and C-terminal fragments of human fibrobl ast caldesmon expressed in Escherichia coli to tropomyosin and its CNB r fragments. The N-terminal fragment, CaD40 (residues 1-152), binds tr opomyosin, but the interaction is mostly abolished in the presence of actin. CaD40 binds strongly to Cn1B(142-281) of tropomyosin, but weakl y to Cn1A(11-127). The C-terminal fragment, CaD39, which corresponds t o residues 443-736 of gizzard caldesmon, binds tropomyosin, and the in teraction is enhanced by actin. CaD39 binds to both Cn1A(11-127) and C n1B(142-281) of tropomyosin. Our results suggest that the N-terminal d omain of caldesmon interacts with the C-terminal half of one tropomyos in molecule, whereas the C terminal domain binds to both N- and C-term inal regions of the adjacent tropomyosin molecule along the actin fila ment. In addition, the binding of the N-terminal domain of caldesmon t o the actin-tropomyosin filament is weak, which may allow this domain to project off the thin filament to interact with myosin.