Fig. 2.

FRET between a donor-labeled FKBP and acceptor attached within CaM's N lobe. (A) Representative spectra in 30 nM Ca²⁺. Samples contained 0 (blue), 100 nM (green), 300 nM (yellow), or 800 nM (red) F-CaM acceptor. The peak at 520 nm reflects F-FKBP fluorescence (excitation at 490 nm). Peak at 600 nm is F-CaM acceptor. Dashed gray line indicates fluorescence of a sample containing 800 nM F-CaM plus 16 μM unlabeled CaM. (B) FRET is plotted as a function of F-CaM acceptor concentration (means ± SEM from 3 experiments). (C and D) FRET between the F-FKBP donor and a Ca²⁺-insensitive F-CaM (F-CaM₁₂₃₄). Representative spectra were obtained in 30 nM Ca²⁺ and either 0, 100, 300, or 800 nM F-CaM₁₂₃₄. Average data are from 3–4 experiments using the F-CaM₁₂₃₄.