Anda Trifan, University of Illinois at Urbana-Champaign
Ras proteins are small GTPases involved in key cell-signaling pathways regulating cell growth, proliferation and division. Overactivity of Ras proteins has been a hallmark of diverse forms of cancer in humans. Interconversion between the active (GTP-bound) and inactive (GDP-bound) forms is mediated by two key protein partners that bind and interact with Ras on the surface of the membrane: hydrolysis of GTP, and inactivation of Ras, is mediated by GAP, while exchange of GDP by GTP is facilitated by GEF, reactivating Ras. While the G-domain, the globular domain that hydrolyzes GTP, of Ras isoforms is highly conserved sequentially and structurally, the difference lies in the hypervariable region (HVR). The HVR plays a crucial role in anchoring the G-domain into the cytosolic membrane. We used K-Ras as it is the most common isoform whose mutations lead to a number of cancer type. The orientation of the globular domain on the membrane is essential to its ability to interact with downstream effectors. We simulated the GTP- and GDP-bound domains on HMMM membrane composed of 70:30 PS/PC lipids. The G-domain initial orientation in solution was varied by 25 to 30 degrees with respect to the membrane to avoid convergence of results due to initial placement bias. While the differences in structure are small in the GTP- and GDP-bound domains, they bind different effectors, therefore their individual orientation on the membrane is important. Our simulations of more than 3 microseconds show the G-domain, in the absence of the linker samples many conformations on the surface of the membrane, contrasting results from simulations where the protein was bound to the membrane by the HVR, converging to two distinct binding modes. Our findings suggest the linker highly constrains the orientations available for the G-domain to sample.
Abstract Author(s): Anda Trifan, Emad Tajkhorshid