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Simulated Thermal Unfolding of the von Willebrand Factor A Domains

Abstract

The A1 and A2 domains of von Willebrand factor (VWF) have important functions: A1 contains a binding site for platelet glycoprotein Ib (GPIb) while A2 contains a cryptic proteolytic site for the VWF-cleavage enzyme, A Disintegrin And Metalloprotease with a ThromboSpondin type 1 motifs 13 (ADAMTS-13). Because the proteolytic site is fully buried in the native A2 structure, A2 needs to be unfolded to expose its proteolytic site for ADAMTS-13 cleavage. To study the unfolding mechanism of the VWF A domains, we used molecular dynamics (MD) to simulate in atomic details the thermal unfolding of A1 and A2 at high temperatures. The thermal unfolding of A1 and A2 appears very different from their unfolding by tensile forces. At 500 K, unfolding of the central β sheet of A2 starts from the two edges and propagates into the center. β4 and β5 in the center are structurally the most stable and unfolded the latest. However, A2 could be unfolded along different pathways and the unfolded A2 structure is highly flexible. By comparison, A1 is unfolded slower than A2 at 500 K. In even longer time, the unfolding of A1 is limited to the edges of the central β sheet, suggesting a protective role of the N–C terminal disulfide bond.

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Acknowledgments

We thank Dr. Stephen Harvey for providing computational resources for the MD simulations. We also acknowledge NSF TeraGrid for providing supercomputer time via NCSA DAC grant MCB080011N and LRAC grant MCA08X014. This work is supported by NIH grants HL093723 and HL091020.

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Correspondence to Cheng Zhu.

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Associate Editor Edward Guo oversaw the review of this article.

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Figure S1

Comparison between the A2 crystal structure and the A2 homology model. (a) Structural alignment of the A2 homology model (yellow) to the A2 crystal structure (red). The biggest difference in the α4–β4 region is indicated. (b) RMSD of Cα atoms of residue 1496–1669 between the two A2 structures. The largest RMSD in the α4–β4 region is indicated (TIFF 407 kb)

Figure S2

SASAs of the A2 proteolytic site (Tyr1605 and Met1606) during the A2 equilibration at 300 K. Results of both backbones (black curve) and sidechains (gray curve) are shown for simulations with the A2 crystal structure (a) and the A2 homology model (b) (TIFF 421 kb)

Video 1

Thermal unfolding of the A2 crystal structure in the NVE ensemble at 500 K. A2 is oriented with β strands β3, β2, β1, β4, β5, and β6 from the left to the right. A2 structures are colored according secondary structures: α helix in purple, 310 helix in blue, π helix in red, β sheet in yellow, β bridge in orange, turn in cyan, and loop in white. The blue and the red spheres indicate the N- and C-terminal Cα atoms, respectively. The backbone atoms of Tyr1605 and Met1606 adjacent to the proteolytic site are shown as green spheres (AVI 27486 kb)

Video 2

Thermal unfolding of the A2 homology model in the NVE ensemble at 500 K. Representations of A2 structures are the same as Video 1 (AVI 25632 kb)

Video 3

Thermal unfolding of the A2 homology model in the NPT ensemble at 500 K. Representations of A2 structures are the same as Video 1 (AVI 29154 kb)

Video 4

Thermal unfolding of A1 in the NVE ensemble at 500 K. A1 is oriented with β strands β6, β5, β4, β1, β2, and β3 from the left to the right. A1 structures are colored according secondary structures: α helix in purple, 310 helix in blue, π helix in red, β sheet in yellow, β bridge in orange, turn in cyan, and loop in white. The blue and the red spheres indicate the N- and C-terminal Cα atoms, respectively. The N–C terminal disulfide bond is shown as green sticks (AVI 28868 kb)

Video 5

Thermal unfolding of A1 in the NPT ensemble at 500 K. Representations of A1 structures are the same as Video 4 (AVI 13656 kb)

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Chen, W., Lou, J. & Zhu, C. Simulated Thermal Unfolding of the von Willebrand Factor A Domains. Cel. Mol. Bioeng. 3, 117–127 (2010). https://doi.org/10.1007/s12195-010-0117-z

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Keywords

  • Molecular dynamics
  • Thermal unfolding
  • von Willebrand factor
  • A domains