Biological Membranes - A. N. Ogurtsov 2012

Structure and Functions of Biomembranes
Intermolecular Interactions in Biomembranes
Lipid-Protein Clusters in Membranes

Recent studies have disproved the long-held view that the lipid monolayer is simply a random mixture of Lipids distributed uniformly throughout it. It was initially discovered that detergent extraction of Plasma Membranes leaves behind a residue enriched in Cholesterol and sphingomyelin. Because these two lipid types form more ordered and viscous bilayers, it was hypothesized that they form microdomains—termed "lipid rafts" or "islands"—surrounded by more fluid phospholipid regions that are preferentially extracted by detergents.

Subsequent experiments have confirmed this hypothesis. For instance, Fluorescence Cell/15.html">Microscopy has revealed the aggregation of lipids and Proteins into such membrane "islands."

Figure 33 illustrates an experimental setup in which The addition of fluorescently labeled cholera toxin molecules (gray diamonds "1") and Antibodies (Y-shaped molecules "2") confirmed, via fluorescence microscopy, that GM1 glycosphingolipids and lipid-anchored Membrane Proteins—specifically placental alkaline phosphatase (PLAP)—co-assemble into lipid-protein "islands" within the membrane.

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Figure 33. Fusion of membrane islands induced by cholera toxin molecules (1) and anti-PLAP antibodies (2)

Cholera toxin forms multiple cross-links with closely spaced GM1 molecules, while antibodies specific to PLAP can similarly bind to two adjacent protein molecules. Consequently, Treatment with these toxins and antibodies leads to The formation of patches composed of Glycolipids and PLAP proteins.

The cholera toxin molecules were labeled with green fluorescent markers, whereas the antibody molecules were labeled in red. If GM1 and PLAP were not part of the same "islands," the resulting patches formed by the action of cholera toxin and antibodies would appear in the Microscope as distinct green or red fluorescent spots, respectively. However, the experiment predominantly revealed yellow spots (yellow being the combination of red and green). This demonstrates that GM1 and PLAP were originally constituents of the same lipid-protein clusters (islands), and the application of toxin and antibodies caused these islands to coalesce.

A different picture emerges in membranes containing transferrin receptors (TfR) instead of PLAP (Figure 34).

Figure 34. Aggregation of GM1 glycosphingolipids and transferrin receptors (TfR) upon treatment with cholera toxin molecules (1) and anti-PLAP antibodies (2), respectively

Treatment of such membranes with the same fluorescently labeled cholera toxin (green) and anti-TfR antibodies (red) resulted in predominantly green and red patches within the fluorescence microscope field of view. Consequently, in this case, GM1 glycosphingolipid molecules formed their own (green) patches under METABOLISM/18.html">The Influence of cholera toxin, while TfR proteins formed separate (red) patches upon antibody binding. This proved the absence of TfR proteins from these lipid-protein clusters.

Lipid-protein islands vary in size, but typically have a diameter of ~50 nm. They can be disrupted by methyl-$eta$-cyclodextrin, which depletes membrane cholesterol, or by Antibiotics such as filipin, which binds cholesterol. This underscores the critical role of cholesterol in driving the aggregation of lipids and proteins into clustered islands.

In addition to cholesterol and Sphingolipids, these islands incorporate numerous cell Membrane Receptors as well as a variety of signaling proteins that bind to and are activated by these receptors. Such lipid-Structure/178.html">Protein Complexes can only form within the two-dimensional environment of the hydrophobic bilayer, and they are thought to facilitate the detection of extracellular chemical signals and the activation of appropriate intracellular responses.

So-called lipid-protein and Protein-Protein Interactions play a crucial role in the structural stabilization of lipid islands. These terms encompass a wide range of phenomena, differing in mechanism, that lead to the non-uniform distribution of molecular components within membranes—a property known as membrane microheterogeneity.