Visualizing RPE65 Peptide-Membrane Interactions Using the Energy Landscape Visualization Method (ELViM)

Amphipathic α-helices (AHs) in eukaryotic, viral, and bacterial proteins mediate diverse functions beyond membrane anchoring, including lipid sensing, curvature detection, and membrane remodeling. Despite advances in structural biology, the conformational dynamics of peripheral membrane proteins (PMPs) during membrane binding remain poorly understood ¹.

Here, we investigate retinol isomerase RPE65, a PMP critical for the vertebrate visual cycle, which contains an AH (residues 107–125) that transitions between a disordered loop and a membrane-binding helix. We aim to elucidate how structural variations in this AH influence membrane affinity by comparing the wild-type (WT) RPE65 peptide to four mutants (C112Y, N114H, R118S [pathogenic]; R124Q, F119L [non-pathogenic]) from the Leiden Open Variation Database (LOVD).

Using molecular dynamics simulations, we modeled the interaction of each peptide (initially folded) with a DOPC/C7DHP (3:1) bilayer membrane. Structural and folding behaviors were analyzed via the Energy Landscape Visualization Method (ELViM)², revealing key differences between pathogenic and non-pathogenic variants. Our preliminary results suggest that the n-terminal region helicity is critical in determining AH pathogenicity; non-pathogenic mutations and the wild type (WT) present more stable α-helical conformations, while pathogenic mutants present more disordered (non-helical) n-terminal conformations.