Molecular Modeling and Solution NMR Studies of the Structure and Dynamics of K-Ras at a Lipid Membrane Containing PIP2

Abstract

Due to its vital role in regulating the cellular signal transduction and a high occurrence in human tumors, the small GTPase K-Ras has been the object of intense experimental and computational studies recently. In particular, research has focused on the role of membrane anchor on the structure and function of K-Ras. In this research, we investigate the structure and dynamics of K-Ras (GTP bound state) interacting with lipid membranes containing PIP2 (Phosphatidylinositol 4, 5-bisphosphate) by performing long time scale all atom molecular dynamics simulation with the CHAMM36 force field. The simulations show that that the K-Ras catalytic domain binds to the membrane within 100 ns, mainly due to the electrostatic attraction between membrane and K-Ras catalytic domain. PIP2 lipids, which possess four negative charges, form persistent (up to several hundreds of nanoseconds) contacts with residues (Arginine and Lysine) from the GTPase catalytic domain. Further, the lipid bound K-Ras presents several different membrane orientations with respect to the membrane. We find that the competitive interaction between the attraction of membrane with the positively charged protein residues and repulsion of membrane with the negatively charged residues likely determines the K-Ras4A orientation, but this also influenced by the extent of the K-Ras surfaces (in relation to the proteins shape)The simulation results are validated by an NMR solution study of K-Ras with nanodiscs and liposomes. Both the full length and hypervariable region truncated GTPase interact transiently with PIP2 doped membrane model systems through its catalytic domain. Similarly to another experimental and simulation study of K-Ras4B (reviewed by us in [1]), the results imply that membrane orientations could be correlated with K-Ras function by hindering or exposing the effector binding domain of the GTPase to solvent. Furthermore, the effect of membrane binding on the flexibility and conformation of nucleotide binding region, especially with the switch loops of K-Ras will be discussed. Reference: [1] Li, Z. L.; Cao S.; Buck, M.; New & Notable: K-RAS at Anionic Membrane: Orientation, Orientation⋯Orientation. Recent Simulations and Experiments. Biophys. J. 2016, 5, 1033-1035.