Mechanisms Of Membrane Binding Of K-Ras4b Farnesylated Hypervariable Region

K-Ras4B belongs to a family of small GTPases that regulates cell growth, differentiation and survival. Kras is frequently mutated in cancer. K-Ras4B modulates downstream signaling at different lipid microdomains. K-Ras4B association with the plasma membrane through its farnesylated and positively charged C-terminal hypervariable region (HVR) is critical to its oncogenic function. HVR farnesylation plays an important role in Ras signaling, but the structural mechanisms and consequences of farnesyl-induced membrane binding are poorly understood and the basis for Ras localization to certain parts of plasma membrane is unknown. Application of confocal microscopy, surface plasmon resonance, and molecular dynamics (MD) simulations revealed that K-Ras4B is distributed between rigid lipid rafts and disordered non-raft regions of the plasma membrane. Its membrane binding domain interaction with phospholipids is driven by membrane fluidity. The farnesyl group spontaneously inserts into the disordered lipid microdomains containing unsaturated phospholipids, while the rigid microdomains composed of tightly packed saturated lipids restrict the farnesyl group penetration. We speculate that the resulting farnesyl protrusion towards the cell interior allows oligomerization of K-Ras4B membrane binding domain in rigid microdomains. Unlike other Ras isoforms, K-Ras4B HVR contains a single farnesyl modification and positively charged polylysine sequence. The high positive charge not only modulates specific HVR binding to anionic phospholipids, but the farnesyl9s membrane orientation. Phosphorylation of Ser181 prohibits spontaneous farnesyl membrane insertion. We speculate that if the farnesyl group is not inserted into the membrane, it may serve in cooperative binding of other K-Ras4B molecules, thus leading to dimerization. In this case, the protein binds the membrane using electrostatic interaction between its positively charged HVR and the negatively charged membrane phospholipids. In the case of disordered phospholipid domains, the farnesyl group burial in the phospholipids tends to hinder its interactions. The mechanism illuminates the roles of HVR modifications in K-Ras4B targeting to microdomains of the plasma membrane and suggests an additional function for HVR in regulation of Ras signaling. Elucidating the mechanism for K-Ras4B localization at different membrane microdomains is important for in-depth understanding of Ras-effector interactions mediating signaling pathways. Funded by Frederick National Laboratory for Cancer Research, National Institutes of Health, under contract HHSN261200800001E. Citation Format: Hyunbum Jang, Sherwin J. Abraham, Tanmay S. Chavan, Ben Hitchinson, Lyuba Khavrutskii, Nadya I. Tarasova, Ruth Nussinov, Vadim Gaponenko. Mechanisms of membrane binding of K-Ras4B farnesylated hypervariable region. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2457. doi:10.1158/1538-7445.AM2015-2457