Fig 1: Detailed BMP10:BMPRII interface interactions.a BMP10:BMPRII binding interface in complex AC. BMP10 (in coral)-binding surface on BMPRII (purple) can be broadly divided into three regions. The central hydrophobic triad (Y67, W85 and F115), the β4 strand to the A-loop (green oval), the F3-loop and the region connecting the A-loop and the F3-loop (light blue circle and orange oval). b–f Detailed interactions between BMP10 and BMPRII, at the β4 strand and A-loop region in all complexes (b), at the F3-loop (c) and the regions connecting the A-loop and the F3-loop in complex AC (d), at BMPRII S107 region in complex BD (e) and AB (f). BMP10 is shown in coral sticks throughout b–f, whereas BMPRII is in grey for complex BD, in dark blue for complex AB, in yellow for complex DL, in magenta for complex CK, in cyan for complex BJ and in green for complex AI. In a–f red dashed lines denote H-bonds, with distance all between 2.7-3.7 Å if not labelled. Underlined residue numbers are those from BMP10, and residues numbers in normal text are those from BMPRII. g Buried interface area between BMP10 and BMPRII in binary and ternary receptor complexes. *Complex BD has much smaller buried surface area because some sidechains at the interface were deleted between the A-loop and F3-loop due to poor densities.
Fig 2: Context-dependent fingertip 3/4 conformation in BMP9 and BMP10.a An overlay of all BMP10 monomers from different protein interaction contexts. Sources of structures are listed in Supplementary Table 5. b A close-up view of fingertip 3/4 with all the structures annotated. c A closed-up view of fingertip 3/4 region after superposition of all BMP9 monomers from different protein interaction contexts. Sources of the structures are listed in Supplementary Table 5. The overlay of the full monomers can be found in Supplementary Fig 9. d A schematic diagram depicting the context-dependent conformation of fingertip 3/4 in BMP9 and BMP10.
Fig 3: BMP10 binding to BMPRII and its mutant proteins.a The sensorgram of monomeric BMPRII WT ECD binding to BMP10 immobilised on a CM5 Biacore chip. Control experiments of ALK1-Fc and BMPRII-Fc binding to the same BMP10 chip are shown in Supplementary Fig 10. b G89-containing A-loop is essential for BMPRII binding to BMP10. c F1-loop and F3-loop deletion mutants binding to BMP10. d Locations of PAH mutations6 on the BMPRII ECD structure (purple cartoon). Red spheres: residues predicted to be deleterious; cyan spheres, residues predicted to be benign. e Sensorgrams of BMPRII ECD proteins containing PAH mutations binding to BMP10. f. Summary of kinetic parameters from the Biacore binding experiments. *G89A has also been found in an hereditary PAH case, but predicted to be a benign mutation. WT wild type, ECD extracellular domain, ΔF1(F3) deletion of finger 1 (finger 3) residues, ΔGDP deletion of Gly, Asp and Pro in the A-loop. M Molar concentration, s = second, ka = association rate constant, kd = dissociation rate constant, KD = kd /ka.
Fig 4: Comparison of ALK1 binding sites in binary and ternary BMPRII receptor complexes.a Overlay of BMP10:ALK1 from the ternary signalling complex (cpx1, magenta, cpx2, purple) to those from binary complexes (PDB code 6SF1 in grey; 6SF3 in cyan for BMP10 and orange for ALK1). The backbones of all overlaid molecules are shown in ribbon. BMP10 from 6SF3 also shown in semi-transparent cartoon. Because in both 6SF1 and 6SF3, there was only one copy of BMP10:ALK1 monomer in an asymmetric unit, the dimeric receptor complexes for 6SF1 and 6SF3 were generated with a symmetry-related molecule and the two BMP10:ALK1 interfaces in 6SF1 and 6SF3 dimer would be identical. b Comparison of the buried surface area at the BMP10 and ALK1 interface in binary and ternary receptor complexes. c Overlay of all ALK1 chains, displayed in ribbon on BMP10 surface (light cyan). Four parts of BMP10 binding sites on ALK1 identified previously13 are highlighted by dashed lines. The colour for each chain is shown below. d Zoomed-in views of ALK1 H87 and E59 interaction area. H-bond interactions are shown with dotted lines. Same colour scheme as in c. The only interactions can be seen are in grey (from 6SF1), and orange and cyan (from 6SF3). Detailed information and the list of the interactions can be found in Supplementary Table 2.
Fig 5: Comparison of BMP10 binding site on BMPRII with BMP9 site on ActRIIB.a Sequence alignment of BMPRII ECD and ActRIIB ECD. Residues at the binding interface with BMP10 or BMP9 are shown in blue. Residues that are not modelled in the BMPRII structure are shown in grey. Hydrophobic triad residues are highlighted in cyan. Positions of the four loops are highlighted below the sequence. Lines above the sequence highlight the residues deleted in the mutagenesis studies. ΔF1(F3) = deletion of finger 1 (finger 3) residues; ΔGDP = deletion of residue Gly, Asp and Pro. b, c BMP10-binding site on BMPRII (b, light purple) and BMP9-binding site on ActRIIB (c, cyan). Residues making direct interactions with BMP10 or BMP9 are coloured in magenta, with hydrophobic triad residues shown in sticks. BMP10 G89 is highlighted in a magenta sphere. Water molecules that mediate hydrogen bond interactions between BMPRII and BMP10 are shown in yellow spheres. d BMPRII in binary (complex AC, purple cartoon) and ternary (complex AI, green cartoon) complexes on BMP10 surface (pale blue), showing G89 (magenta spheres) docking into a deep pocket on BMP10. e ActRIIB (cyan cartoon) on BMP9 surface (pale blue, from PDB entry 4FAO) does not have A-loop mediated interaction. In d, e, residues making H-bond interactions shown in magenta, and hydrophobic triad shown in magenta sticks.
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