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Antibody binding reports spatial heterogeneities in cell membrane organization.

ABSTRACT: The spatial organization of cell membrane glycoproteins and glycolipids is critical for mediating the binding of ligands, receptors, and macromolecules on the plasma membrane. However, we currently do not have the methods to quantify the spatial heterogeneities of macromolecular crowding on live cell surfaces. In this work, we combine experiment and simulation to report crowding heterogeneities on reconstituted membranes and live cell membranes with nanometer spatial resolution. By quantifying the effective binding affinity of IgG monoclonal antibodies to engineered antigen sensors, we discover sharp gradients in crowding within a few nanometers of the crowded membrane surface. Our measurements on human cancer cells support the hypothesis that raft-like membrane domains exclude bulky membrane proteins and glycoproteins. Our facile and high-throughput method to quantify spatial crowding heterogeneities on live cell membranes may facilitate monoclonal antibody design and provide a mechanistic understanding of plasma membrane biophysical organization.

SUBMITTER: Arnold DP

PROVIDER: S-EPMC10199101 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Antibody binding reports spatial heterogeneities in cell membrane organization.

Arnold Daniel P DP Xu Yaxin Y Takatori Sho C SC

Nature communications 20230520 1

The spatial organization of cell membrane glycoproteins and glycolipids is critical for mediating the binding of ligands, receptors, and macromolecules on the plasma membrane. However, we currently do not have the methods to quantify the spatial heterogeneities of macromolecular crowding on live cell surfaces. In this work, we combine experiment and simulation to report crowding heterogeneities on reconstituted membranes and live cell membranes with nanometer spatial resolution. By quantifying t ...[more]

PMID: 37208326

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Project description:The ligands of certain G-protein coupled receptors (GPCRs) are membrane soluble and reach their target from the lipid bilayer. Lipid composition and dynamics will therefore modulate the activity of these receptors, but specific roles of lipid components, including the ubiquitous cholesterol (Chol), are not clear. We have probed the organization and dynamics of such a lipid-bilayer-penetrating ligand, the endogenous ligand for the κ-opioid receptor (KOR) dynorphin A (1-17) (DynA), using molecular dynamics (MD) simulations of DynA in cholesterol-depleted and cholesterol-enriched model membranes. DynA is found to penetrate deep inside fluid dimyristoylphosphatidylcholine (DMPC) bilayers, and resides with its N-terminal helix at ∼6 Å away from the bilayer midplane, in a tilted orientation, at an ∼50° angle with respect to the membrane normal. In contrast, DynA inside DMPC/Chol membranes with 20% cholesterol (DMPC/Chol) is situated with its helical segment ∼5 A higher, i.e., closer to the lipid/water interface and in a relatively vertical orientation. The DMPC membrane shows greater thinning around the insertion and permits a stronger influx of water inside the hydrocarbon core than the DMPC/Chol membranes. Relating these results to data about key GPCR residues that have been implicated in interactions with membrane-inserting GPCR ligands, we conclude that the position of DynA in DMPC/Chol, but not in pure DMPC, correlates with generally proposed GPCR ligand entry pathways. Our predictions provide a possible mechanistic explanation as to why DynA binding to KOR, and the subsequent activation of the receptor, is facilitated in cholesterol-enriched environments. A quantitative description of DynA-induced membrane deformations is obtained with a continuum theory of membrane deformations (CTMD) that is based on hydrophobic matching. Comparison with the MD data reveals the significance of the lipid tail packing energy contribution in the DMPC/Chol mixtures in predicting equilibrium membrane shape around DynA. On this basis, specific corrections are introduced to this energy term within the CTMD framework, thereby extending the applicability of the CTMD framework to lipid raft mixtures and their interactions with GPCR proteins and their ligands.

Dataset Information

Antibody binding reports spatial heterogeneities in cell membrane organization.

Publications

Antibody binding reports spatial heterogeneities in cell membrane organization.

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