Different polymeric forms of actin detected by the fluorescent probe terbium ion.
The interaction of actin with Tb3+ was studied by following the fluorescence emitted at 545 nm when the protein was excited at 285 nm in the presence of Tb3+. It was shown that, at low ionic strength, each actin monomer binds, at saturation, six Tb3+ with an association constant of 0.8 microM-1. In the presence of 0.1 M KCl the association constant decreases to 0.15 and 0.24 microM-1 at subcritical and overcritical actin concentrations, respectively; the number of the binding sites remains six. When polymeric actin is formed by the addition of 2 mM MgCl2, the association constant drops to 0.008 micro M-1 and the number of the binding sites to four. The lower number of the Tb3+ binding sites (four) in the actin polymerized by MgCl2 as compared to the number of binding sites (six) of the actin polymerized by KCl is taken as evidence of a looser structure of this latter polymer. We have also shown that Tb3+ does not replace 45Ca2+ at the single, "high-affinity" site of G-actin. Removal of this Ca2+, in the presence of Tb3+, destroys the typical G- and F-actin structures.
D000199:Actins / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D008433:Mathematics; D009132:Muscles / Q000378:metabolism; D011485:Protein Binding; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D013725:Terbium / Q000494:pharmacology*
D000199:Actins; D046911:Macromolecular Substances; D013725:Terbium; D002118:Calcium