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Nucleotide and actin binding properties of the truncated myosin head (S1dC) from Dictyostelium myosin II were studied in solution using rabbit skeletal myosin subfragment 1 as a reference material. S1dC and subfragment 1 had similar affinities for ADP analogues, ɛADP and TNP-ADP. The complexes of ɛADP and BeFx or AlF4 - were less stable with S1dC than with subfragment 1. Stern-Volmer constants for acrylamide quenching of S1dC complexes with ɛADP, ɛADP·AlF 4 - and ɛADP.BeFx were 2.6, 2.9 and 2.2 M-1, respectively. The corresponding values for subfragment 1 were 2.6, 1.5 and 1.1 M-1. The environment of the nucleotide binding site was probed by using a hydrophobic fluorescent probe, PPBA. PPBA was a competitive inhibitor of S1dC Ca2+-ATPase (Ki = 1.6 μm). The binding of nucleotides to subfragment 1 enhanced PPBA fluorescence and caused blue shifts in the wavelength of its maximum emission in the order: ATP ≈ ADP·AlF4- ≈ ADP·BeFx > ATPSγS > ADP > PPi. In the case of S1dC, the effects of different nucleotides were smaller and indistinguishable from each other. S1dC bound actin tighter than S1 (Kd = 7 nm and 60 nm, respectively). The actin activated MgATPase activity of S1dC varied between preparations, and the Vmax and Km values ranged between 3 and 7 s-1 and 60 and 190 μm, respectively. S1dC showed lower structural stability than S1 as revealed by their thermal inactivations at 35° C. These results show that the nucleotide and actin binding of S1dC and subfragment 1 are similar but there are some differences in nucleotide and phosphate analogue-induced changes and the communication between the nucleotide and actin binding sites in these proteins