Structural vulnerability in EPCR suggests functional modulation

Figures

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  Figure 1. The non-canonical EPCR structure. (A) The canonical (left) and non-canonical EPCR structures are shown in blue and pale yellow colors, respectively. The residues in the α2 helix with severe folding transitions are highlighted. 2Fo-Fc electron density maps are shown as dark grey meshes. The lipid is not shown to facilitate the focus on the vul region (B) Residues Ala153-Tyr154-Asn155 were omitted from the protein coordinates, the structure refined and the Fo-Fc electron density map (green mesh) was calculated to show an unbiased signal of the vul region. (C) Superposition of the canonical and non-canonical structures with Tyr154 highlighted as sticks. The lipid is not shown. (D) Top view of EPCR with the alpha helices shown as cylinders and the structurally vulnerable region (vul region) highlighted in blue. The bound lipid is shown inside the EPCR cavity.

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  Figure 2. Impact of the non-canonical folding mode in protein C/APC binding. (A) Structure of the protein C Gla domain (in orange color) in complex with the canonical conformation of EPCR (PDB 1LQV). The contacts with the Gla domain established by EPCR Tyr154 are highlighted with grey dashed lines. Tyr154 residues in both the canonical and non-canonical EPCR structures are highlighted in sticks for comparison purposes and to better visualize the impact of the folding transition in protein C binding. The bound lipid can be observed inside the pocket as sticks. (B) Top views of Asn155, Arg156 and Thr157 in the vul region. Note that the side chain of Arg156 is omitted due to the lack of electron density (C) Intermolecular contacts (within 4 Å) established between Tyr154 side chain and the N-acetylglucosamine (NAG) present in a crystallographic symmetry mate molecule. (D) A direct and visual comparison of the binding profiles of wt EPCR and EPCR Y154A to 200 nM APC is shown in a biolayer interferometry sensorgram. The full binding kinetics with wt APC is displayed in the lower panel. Black and red lines indicate, respectively, the raw data and the fitting of the association and dissociation…