Biophysical Society Thematic Meeting | Ascona, Switzerland

Liposomes, Exosomes, and Virosomes: From Modeling Complex Membrane Processes to Medical Diagnostics and Drug Delivery

Poster Abstracts

29-POS Board 15 Membrane Curvature Sensing, Membrane Deformation and Lipid Binding Assayed Simultaneously for N-BAR Proteins in Newly Developed Single-Vesicle Assay: FlowSLiC Rasmus Herlo 1,2 , Jannik Larsen 2 , Dimitrios Stamou 2 , Kenneth L. Madsen 1 , Nikolaj Christensen 1 , Ulrik Gether 1 . 1 Department of Neuroscience and Pharmacology, University of Copenhagen, Copenhagen, Denmark, 2 Department of Neuroscience and Pharmacology & Nano-Science center, University of Copenhagen, Copenhagen, Denmark. The expanding sub-group of BAR (Bin/Amphiphysin/Rvs) domain proteins, the N-BARs, contain an N-terminal amphipathic helix (AH). AHs can sense membrane curvature, and have accordingly been identified as the membrane curvature sensing (MCS) segment of N-BARs. The insertion of amphipathic helix into the lipid membranes has also been demonstrated to deform membranes (MemDef), often through low-throughput bulk vesiculation assays, but the interplay between the helix and the scaffolding BAR domain dimer is still subject to debate. In addition, the lipid binding for N-BARs often includes a oligomerization regime, potentially involving an AH-mediated lattice formation. Here we have identified a novel amphipathic helix in PICK1 from the Arfaptin group of BAR proteins, and characterized the lipid interaction regimes through our previously developed Single Liposome Curvature-sensing (SLiC) assay. We show how these parameters control the localization and activity of the PICK1. We thereafter utilize the knowledge of the lipid binding capacities of this protein, as well as the bona fide N-BAR protein Endophilin, to develop a new Flow Cytometry-based assay to explore the interaction between proteins and liposomes. The assay we hence named; FlowSLiC. FlowSLiC is high-throughput, advantageously assaying all three cardinal features of lipid binding (MCS, MemDef, oligomerization) simultaneously. Here, we use it to delineate the concentration- and time-dependent components in the binding regimes of these two N-BARs, but the generality of the assay makes it suited to assay the binding capacities of lipid binding proteins wide across biology.

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