A Computational Study of H-Ras Nanoclusters in Membrane Domains

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The Ras family of enzymes are guanine triphosphatases (GTPases) that function as molecular switches by cycling between GDP-bound off and GTP-bound on conformational states. Malfunction of Ras proteins due to somatic mutations accounts for about 30% of human tumors. The signaling function of Ras proteins is highly related to their ability to form protein-lipid nanodomains (termed nanoclusters) on the plasma membrane. However, the molecular basis for the formation and distribution of Ras nanoclusters has not been determined. We attempt to address this fundamental issue by studying H-Ras proteins in model membranes and focusing on (1) how multiple Ras proteins oligomerize on the membrane surface, (2) how nanoclustering might be affected by conformational variations, and (3) how nanoclusters might affect the host membrane.To achieve these goals, we performed coarse-grained molecular dynamics simulations of full-length GDP- and GTP-bound H-Ras in a model membrane. We found that variation in the initial conformation of these two states of H-ras leads to nanoclusters that exhibit different dynamic behaviors. Analysis of protein-protein contacts in the clustered proteins allowed us to map the residues involved in aggregation. By zooming in on the individual residues involved in protein-protein interactions, we found that the two states of H-ras significantly differ in the accessibility and availability of the structural elements that are required for effector binding. Another important observation is that nanoclusters generate positive curvatures on both layers of the membrane. To investigate the mechanisms of these membrane deformations, we performed three dimensional pressure field analyses and determined the surface tension and elastic bending modulus of each monolayer. These results highlight the intricacies of Ras nanocluster formation, which involve both protein-membrane and protein-protein interactions, and pave the way for a better understanding of signal transduction events mediated by Ras clusters.