PB0820 - In silico evidence of the conformational transitions associated with the p.R854Q mutation in the D’D3 domain of von Willebrand factor, responsible for its weakened FVIII interaction.

Background: The rare type-2N von Willebrand’s disease (2N-VWD) causes in the heterozygous form a mild bleeding diathesis, although homozygous variants are associated with severe bleeding. In homozygous form, 2N-VWD causes in fact a disproportionately low factor VIII (FVIII) level, secondary to a reduced FVIII binding to the D’D3-domain of VWF with consequent increase of its clearance. The VWF-p.R854Q variant is found in almost half of these patients.

Aims: This molecular modeling study aims to analyze the conformational transitions of D’D3 domain associated with the R854Q mutation and the effects on FVIII binding.

Methods: The models of WT-VWF-D’D3 domain (S764-G1241) and its p.R854Q pathological variant were obtained by I-TASSER server using the “6n29” pdb file as a template. The human FVIII-(BDD) structure was derived from the “2r7e” pdb file (A1-A2-A3-C1-C2-domains). Docking was studied with the ClusPro server. Validation of the models was performed with PROCHECK using the PDBsum server. The energetics of D’D3-FVIII binding was calculated with the PRODIGY program, and the models rendering using the Pymol visualization tool.

Results: The R854Q mutation eliminated the polar interactions of R854 with V850, D860, and T859, disrupting the helix formed by T848 and W856, transformed into two antiparallel β-strands. The conformational transition in this region causes an allosteric rearrangement of the entire D’D3 domain (see Fig.1). The latter showed a different orientation in the binding to FVIII in comparison to the WT form, engaging different and much fewer residues than the WT form (7 vs 29, Fig. 2). The energetics of the R854Q-D’D3-FVIII interaction showed a strongly lower affinity than the WT form (ΔΔG≃10 kcal/mol).

Conclusion(s): VWF-R854 does not directly interact with FVIII, although it stabilizes a correct D'D3 conformation for binding to FVIII. The p.R854Q mutation induces a conformational change in FVIII, causing a strong reduction of FVIII affinity.