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. 2005 Feb 8;386(Pt 1):73–83. doi: 10.1042/BJ20041356

Figure 3. Dynamic association/dissociation of purified VDAC as demonstrated with FRET.

Figure 3

VDAC was purified using β-OG and labelled with FITC or EITC. Oligomers labelled by both fluorophores were formed as described under the Experimental section. (A) SDS/PAGE of purified VDAC labelled with FITC (lane 1), EITC (lane 2), a mixture of FITC- and EITC-VDAC (lane 3) and the FITC- and EITC-VDAC mixture containing added unlabelled VDAC (lane 4), followed by Coomassie Blue staining (Coomassie) or exposure to UV light to visualize the fluorescence. (B) The emission spectra of FITC- and EITC-VDAC were measured separately, with their computed sum presented (○). The emission spectrum of a mixture of FITC- and EITC-VDAC (●), and that of the FITC- and EITC-VDAC mixture after subjecting to dissociation–association conditions (——), are presented. FRET is evident by the quenching of the donor (FITC) emission and an increase in acceptor (EITC) emission. (C) Emission spectra of FITC- and EITC-VDAC ‘hetro-oligomers’ formed in the absence (——) or presence of an approx. 2-fold excess of unlabelled VDAC (●). This experiment is representative of four similar experiments. AU, arbitrary units.