Dissociation of the actin.subfragment 1 complex by adenyl-5'-yl imidodiphosphate, ADP, and PPi.
The ability of adenyl-5'-yl imidodiphosphate (AMP-PNP), ADP, and PPi to dissociate the actin.myosin subfragment 1 (S-1) complex was studied using an analytical ultracentrifuge with UV optics, which enabled the direct determination of the dissociated S-1. At mu = 0.22 M, pH 7.0, 22 degrees C, with saturating nucleotide present, ADP weakens the binding of S-1 to actin about 40-fold (K congruent to 10(5) M-1), while both AMP-PNP and PPi weakens the binding about 400-fold (K congruent to 10(4) M-1). This 10-fold stronger dissociating effect of AMP-PNP and PPi compared to ADP correlates with our data showing that the binding of AMP-PNP and PPi to S-1 is about 10-fold stronger than the binding of ADP. In contrast, the binding constants of ADP, AMP-PNP, and PPi to acto.S-1 are nearly identical (K congruent to 5 x 10(3) M-1). At 4 degrees C, AMP-PNP has only a 3-fold stronger dissociating effect than ADP and, similarly, our data suggest that the binding of AMP-PNP and ADP to S-1 is quite similar at 4 degrees C. AMP-PNP and PPi are, therefore, somewhat better dissociating agents than ADP, but the difference among these three ligands is quite small. These data also show that actin and nucleotide bind to separate but interacting sites on S-1 and that the S-1 molecules bind independently along the F-actin filament with a binding constant of about 1 x 10(7) M-1 at 22 degrees C and physiological ionic strength.
D000199:Actins / Q000378:metabolism; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives; D000266:Adenylyl Imidodiphosphate; D000818:Animals; D001665:Binding Sites; D011756:Diphosphates; D007460:Iodoacetamide; D007700:Kinetics; D046911:Macromolecular Substances; D009132:Muscles / Q000378:metabolism; D009218:Myosins / Q000378:metabolism; D011485:Protein Binding; D011817:Rabbits
D000199:Actins; D011756:Diphosphates; D046911:Macromolecular Substances; D000266:Adenylyl Imidodiphosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D009218:Myosins; D007460:Iodoacetamide