Figure 1.
Modeling D′D3 dimerization. (A) Domain architecture and position of dimerizing cysteine residues (red vertical lines with S) in D′D3 assembly (top) and VWF monomer (bottom). (B) Solvent-accessible surface of D′D3.9 (C-D) Sequence alignments of D3 C8-3 and TIL3 modules in VWF and gel-forming mucins (see supplemental Figure 1 for VWD3 and E3). Cysteines implicated in disulfide exchange and interchain disulfides are noted with asterisks (*) and highlighted in red, whereas those that form intrachain disulfides are linked and highlighted in other colors. (E-G) The D′D3 dimer model is shown as a solvent-accessible surface colored by module with its twofold rotational axis as a double-ended arrow colored green on one end and blue on the other. D3 C termini in E3 are marked with large light-blue spheres. Residues mutated in VWF disease that cause loss of VWF binding (type 2M) and no other phenotypes (types 1, 3, 2A, and 2B) are marked with large red spheres. VWF tubules in Weibel-Palade bodies also have a dyad symmetry axis3,4 to which the D′D3 dimer axis must be parallel. VWF biology suggests that the dimer axis end colored green in panels E through G would orient toward the outside rather than the inside of tubules. This side links to A1 and the VWF C terminus and bears VWF disease mutations that specifically abolish factor VIII binding,9 which occurs during biosynthesis.19 A model of VWF tubules based on MUC2 filaments also supports this orientation.20 The dimer model was created with Modeler 9.621 as described in the text with each monomer templated with the segments of VWF and MUC2 (PDB ID 6N29 and 6RBF, respectively) shown in supplemental Figure 2. (H) Cross-correlation of the D′D3 dimer model to electron microscopy class averages of D′D3-A1 dimers12 was as described.12,22 The best-correlating D′D3 dimer orientation is shown enlarged as a ribbon diagram with coloring similar to that in panels E through G and with spheres at C termini of E3 to show where the mucin linker to A1 attaches. Owing to flexibility of this linker, A1 (a round globule) appears in different orientations or does not appear in class averages. CK, C-terminal cystine-knot domain. Modified from Springer.5

Modeling D′D3 dimerization. (A) Domain architecture and position of dimerizing cysteine residues (red vertical lines with S) in D′D3 assembly (top) and VWF monomer (bottom). (B) Solvent-accessible surface of D′D3.9  (C-D) Sequence alignments of D3 C8-3 and TIL3 modules in VWF and gel-forming mucins (see supplemental Figure 1 for VWD3 and E3). Cysteines implicated in disulfide exchange and interchain disulfides are noted with asterisks (*) and highlighted in red, whereas those that form intrachain disulfides are linked and highlighted in other colors. (E-G) The D′D3 dimer model is shown as a solvent-accessible surface colored by module with its twofold rotational axis as a double-ended arrow colored green on one end and blue on the other. D3 C termini in E3 are marked with large light-blue spheres. Residues mutated in VWF disease that cause loss of VWF binding (type 2M) and no other phenotypes (types 1, 3, 2A, and 2B) are marked with large red spheres. VWF tubules in Weibel-Palade bodies also have a dyad symmetry axis3,4  to which the D′D3 dimer axis must be parallel. VWF biology suggests that the dimer axis end colored green in panels E through G would orient toward the outside rather than the inside of tubules. This side links to A1 and the VWF C terminus and bears VWF disease mutations that specifically abolish factor VIII binding,9  which occurs during biosynthesis.19  A model of VWF tubules based on MUC2 filaments also supports this orientation.20  The dimer model was created with Modeler 9.621  as described in the text with each monomer templated with the segments of VWF and MUC2 (PDB ID 6N29 and 6RBF, respectively) shown in supplemental Figure 2. (H) Cross-correlation of the D′D3 dimer model to electron microscopy class averages of D′D3-A1 dimers12  was as described.12,22  The best-correlating D′D3 dimer orientation is shown enlarged as a ribbon diagram with coloring similar to that in panels E through G and with spheres at C termini of E3 to show where the mucin linker to A1 attaches. Owing to flexibility of this linker, A1 (a round globule) appears in different orientations or does not appear in class averages. CK, C-terminal cystine-knot domain. Modified from Springer.5 

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