Biophysical Society Thematic Meeting

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

, Dimitrios Stamou

, Kenneth L. Madsen

, Nikolaj

Christensen

, Ulrik Gether

Department of Neuroscience and Pharmacology, University of Copenhagen, Copenhagen,

Denmark,

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.