Fig. 4. Fluorescence lifetime measurements fitted taking the instrument response function (IRF) into account by a bi- or tri-exponential decay model for the respective samples. (a) GFP^K25C, apoCyt b₅₆₂^N80C–GFP^K25C, reconstituted dimer obtained by incorporation of heme into apoCyt b₅₆₂^N80C–GFP^K25C, and Cyt b₅₆₂^N80C–GFP^K25C. (b) GFP^K25C and hetero-dimers with GFP^K25C. (c) GFP^S174C and hetero-dimers with GFP^S174C. Conditions: [protein] = 20 μM, 1 mL, prepared in 0.1 M potassium phosphate buffer, pH 7.0 at 25 °C. Full scale fluorescence lifetime measurements are shown in Fig. S10.†.