Calcium-dependent changes in otoferlin fluorescence. (A and B) Normalized plots of the emission intensities of 1 µM of the three–C2 domain fragments C2ABC and C2DEF (A) and 1 µM of the isolated C2 domains C2B, C2C, C2D, C2E, and C2F (B) as a function of increasing calcium concentration in the absence of liposomes. (C and D) Calcium titrations of C2ABC, C2DEF (C), and each isolated C2 domain of otoferlin (D) in the presence of liposomes composed of 15% PS, 55% PC, and 30% PE. Error bars indicate mean ± SD (n = 4).