Figure 5.

LC3B C terminus remains exposed to the hydrophilic environment after protein binding to CL-enriched membranes. Representative NBD fluorescence emission spectra of (A) LC3B^Q15C-NBD (1 µM) in the absence or presence of increasing amounts of liposomes containing either PC:DOPE (80:20 mol ratio), PC:DOPE:CL or PC:DOPE: PtdIns (50:20:30 mol ratio); and (B) LC3B^S101C-NBD (1 μM) in the absence or presence of PC:DOPE:CL liposomes. In each case, fluorescence was normalized to the peak intensity of the protein spectrum in the absence of liposomes. (C) Structural model generated with PyMol depicting the 2 LC3B residues that were individually mutated to cysteine obtaining single-cysteine LC3B mutants. The environmentally sensitive fluorophore NBD was used to label each of these single cysteine residues. PDB: 1UGM. OMM, outer mitochondrial membrane. Norm., normalized.