Flexible-Body Motions of Calmodulin and the Farnesylated Hypervariable Region Yield a High-Affinity Interaction Enabling K-Ras4B Membrane Extraction.

In calmodulin (CaM)-rich environments, oncogenic KRAS plays a critical role in adenocarcinomas by promoting PI3K/Akt signaling. We previously proposed that at elevated calcium levels in cancer, CaM recruits PI3Ka to the membrane and extracts K-Ras4B from the membrane, organizing a K-Ras4B/CaM/PI3Ka ternary complex. CaM can thereby replace a missing receptor tyrosine kinase signal to fully activate PI3Ka. Recent experimental data show that CaM selectively promotes K-Ras signaling, but not of N-Ras or H-Ras. How CaM specifically targets K-Ras and how it extracts it from the membrane in KRAS-driven cancer is unclear. Obtaining detailed structural information for a CaM/K-Ras complex is still challenging. Here, using molecular dynamics simulations and fluorescence experiments, we observed that CaM preferentially binds unfolded K-Ras4B hypervariable regions (HVRs) and not a-helical HVRs. The interaction involved all three CaM domains including the central linker and both lobes. CaM specifically targeted the highly polybasic anchor region of the K-Ras4B HVR that stably wraps around CaM's acidic linker. The docking of the farnesyl group to the hydrophobic pockets located at both CaM lobes further enhanced CaM/HVR complex stability. Both CaM and K-Ras4B HVR are highly flexible molecules, suggesting that their interactions permit highly dynamic flexible-body motions. We therefore anticipate that the flexible-body interaction is required to extract K-Ras4B from the membrane, since conformational plasticity enables CaM to orient efficiently to the polybasic HVR anchor, which is partially diffused into the liquid-phase membrane. Our structural model of the CaM/K-Ras4B HVR association provides plausible clues to CaM's regulatory action in PI3Ka activation involving the ternary complex in cell proliferation signaling by oncogenic K-Ras. Copyright © 2017, The American Society for Biochemistry and Molecular Biology.

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