S100A1 is a typical representative of a group of EF-hand calcium-binding proteins known as the S100 family. The protein is composed of twoasub-units, each containing two calcium-binding loops (N and C). At physiologi-cal pH () and NaCl concentration (100 mm), we determined the microscopic binding constants of calcium to S100A1 by analysing the Ca 2+ -titration curves of Trp90 fluorescence for both the native protein and its Glu32fiGln mutant with an inactive N-loop.