Dorogi_1981_Biophys.Chem_14_91

Stoichiometries, equilibrium constants and optical extinction coefficients of calcium-antipyrylazo III (An) complexing are determined with the analytical method described in article I of this series. Spectrophotometric Ca titrations of An at the wavelengths 595 and 710 nm indicate overall dissociation equilibrium constants for the complexes CaAn, CaAn2 and Ca2An to be 4.5 x 10(-4) M, 1.1 x 10(-8) M2 and 1.5 x 10(-6) M2, respectively, extrapolated to zero ionic strength. Ca titrations of solutions containing An plus acetylcholine receptor protein give clear evidence that An binds to the protein to a large extent in the presence of Ca2+; furthermore, addition of acetylcholine results in release of protein-bound Ca and An. This is the first reported indication that antipyrylazo III binds to biological material and questions the usefulness of this dye as a Ca indicator in biological systems.