1. Raf-1 Cysteine-Rich Domain Increases the Affinity of K-Ras/Raf at the Membrane, Promoting MAPK Signaling 
Shuai Li, Hyunbum Jang, Jian Zhang, Ruth Nussinov 
Structure 
Volume 26, Issue 3, Pages e2 (March 2018)
DOI: /j.str
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2. Structure 2018 26, 513-525.e2DOI: (10.1016/j.str.2018.01.011)
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3. Figure 1 Sequence and Domain Structures of Raf-1
(A) Raf-1 is composed of RBD (residues 56–131), CRD (138–184), and kinase domain (349–609). A long flexible linker connecting the CRD and kinase domain is largely disordered. In the sequence, hydrophobic, polar/glycine, positively charged, and negatively charged residues are colored black, green, blue, and red, respectively. Gray denotes the unstructured region.
(B) Crystal structure of Raf-1 RBD (PDB: 4G0N) and solution structure of Raf-1 CRD (PDB: 1FAR). The structure of 1FAR is a minimized average. Similarly, in the ribbon representation for the secondary structures, the same colors were used, except for the hydrophobic residues, which are colored white.
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4. Figure 2 Membrane Interaction of Raf-1 CRD
(A) Probability distribution functions of the angle, θZn, between the vector connecting two zinc ions in Raf-1 CRD and the direction of membrane normal for eight different CRD configurations (Config. 1–8) interacting with the anionic bilayer composed of DOPC:DOPS (4:1, mole ratio).
(B) Snapshots of Raf-1 CRD interacting with the anionic bilayer depicted from the final conformation at 200 ns. In the bilayer structure, DOPC and DOPS are shown as white and gray surfaces, respectively. In the CRD structure, blues spheres denote zinc ions, and red arrows represent the vector. Key basic residues are marked.
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5. Figure 3 Key Basic Residues of Raf-1 CRD
(A) The probability of lipid contact for each basic residue in Raf-1 CRD at the anionic bilayer composed of DOPC:DOPS (4:1, mole ratio).
(B) Cumulative frequency of the lipid contact for the key basic residues from all configurations.
(C) Binding energy of Raf-1 CRD with the anionic bilayer calculated by the MM-GBSA method for eight different CRD configurations (1–8).
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6. Figure 4 Membrane Interaction of Raf-1 RBD-CRD in Complex with K-Ras4B
Snapshots of GTP-bound K-Ras4B interacting with Raf-1 RBD-CRD at the anionic bilayer composed of DOPC:DOPS (4:1, mole ratio) depicted from the final conformation at 300 ns. In each dimeric configuration (DC), the K-Ras4B-GTP is shown on the left-hand side. In the K-Ras4B cartoon, thick deep-blue tube, yellow, red sticks, and green sphere represent the HVR, farnesyl, GTP, and Mg2+, respectively. The Raf-1 is shown on the right-hand side. The pink and green cartoons are RBD and CRD, respectively. Blue spheres in CRD denote Zn2+. In the bilayer structure, DOPC and DOPS are shown as white and gray surfaces, respectively.
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7. Figure 5 Probability Distribution Across the Lipid Bilayer
Probability distribution functions for different component groups of lipids (CH3, methyl group, red; PO4, phosphate group, orange), different domains of K-Ras4B (catalytic domain, yellow; HVR, green; farnesyl, blue), and different domains of Raf-1 (RBD, dark blue; CRD, purple) as a function of distance from the bilayer center for seven different DCs (1–7) of K-Ras4B/RBD-CRD complex at the anionic bilayer composed of DOPC:DOPS (4:1, mole ratio).
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8. Figure 6 Lipid Contact Probability
The probability of lipid contact for the residues of K-Ras4B (upper), Raf-1 RBD (middle), and Raf-1 CRD (lower) for seven different DCs (1–7) of K-Ras4B/RBD-CRD complex at the anionic bilayer composed of DOPC:DOPS (4:1, mole ratio). Inset in the upper panel highlights the probability for the residues of K-Ras4B in the range of 40–110.
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9. Figure 7 Helix Tilt Angle of K-Ras4B
Probability distribution functions of the helix tilt, θ, with respect to the bilayer normal for two effector lobe helices (α1 and α2) and for three allosteric lobe helices (α3, α4, and α5) in the catalytic domain of K-Ras4B in complex with Raf-1 RBD-CRD for seven different DCs (1–7) at the anionic bilayer composed of DOPC:DOPS (4:1, mole ratio). The helix tilt was calculated for the angle between two vectors formed by the helix axis and normal axis of the bilayer.
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10. Figure 8 K-Ras4B Orientation at the Membrane
Two-dimensional probability distributions of orientation angles for two vectors, CO→ and PO→, in GTP with respect to the bilayer normal sampled from all DCs of K-Ras4B/RBD-CRD complex at the anionic bilayer composed of DOPC:DOPS (4:1, mole ratio). CO→ and PO→ denote the vectors connecting the atom pairs C6 → O6 and PA → O3A in GTP, respectively. The probability surface can be calculated from the occupancy frequency of visiting each grid point on the plane of CO→ and PO→ orientation angles, θCO and θPO, respectively, with respect to the bilayer normal (right). Six K-Ras4B conformations are depicted from the population map, representing the populated K-Ras4B membrane orientations in complex with Raf-1 RBD-CRD. For the protein structures on the membrane surface, the curved arrows on the top of the thick gray arrows measure the molecular inclination to the left- or right-hand sides. The curved arrows on the left measure the molecular rotation at the axis perpendicular to the thick gray arrows representing the molecular inclination to the forward or backward directions.
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