STEADY-STATE AND TIME-RESOLVED FLUORESCENCE MEASUREMENTS FOR STUDYINGMOLECULAR-INTERACTIONS - INTERACTION OF A CALCIUM-BINDING PROBE WITH PROTEINS

The binding of 2-[(2-bis-[carboxymethyl] hylphenoxy)-methyl]-6-methoxy -8-bis[carboxymethyl] aminoquinoline, the fluorescent calcium probe Qu in2, to serum albumin and several other proteins has been investigated . Changes in fluorescence emission spectra and fluorescence anisotropy revealed interactions between Quin2 and several proteins including hu man serum albumin, bovine serum albumin, aldolase, phosphoglucose isom erase, glyceraldehyde-3-phosphate dehydrogenase, and alkaline phosphat ase. Protein-probe interactions were inhibited by the presence of calc ium. Binding was also measured by resonance energy transfer and gel pe rmeation chromatography. Equilibrium binding constants for Quin2 were quantitated by the application of the recently-developed 'SPECTRABIND' program to spectroscopic data (D. Toptygin and L. Brand, Anal. Bioche m., 224 (1995) 330-338). Binding of Quin2 to human serum albumin is di scussed in terms of the published X-ray crystal structure of human ser um albumin (X.M. He and D.C. Carter, Nature, 358 (1992) 209-215).