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Blood, Vol. 108, Issue 12, 3753-3756, December 1, 2006
A single high-affinity binding site for von Willebrand factor in collagen III, identified using synthetic triple-helical peptides
Blood Lisman et al.
108: 3753
Supplemental materials for: Lisman et al, Vol 108, Issue 12, 3753-3756
Files in this Data Supplement:
- Table S1. Sequences of the Collagen III Toolkit (PDF, 37.9 KB) -
Table S1 shows the peptide sequences of the Collagen III Toolkit. Predicted mass is shown, along with melting temperature determined by polarimetry. The deduced VWF-binding motif is highlighted in boldface.
- Table S2. Amino acid sequence of truncated and alanine-modified peptide based on the sequence of peptide no. 23 (top sequence) (PDF, 16.4 KB) -
Triple helical peptides were synthesized with the amino acid sequence shown flanked at the amino terminus by the sequence GPC-(GPP)5- and at the carboxy terminus by -(GPP)5-GPC-NH2 to induce triple helical stability. Peptide sequences are denoted by standard one-letter amino acid nomenclature, in which O represents hydroxyproline. Peptide 2 of this set contains the full VWF-binding sequence, and peptides 7 to 14 represent an alanine scan (substituted A is in boldface) defining the critical residues within the motif. Corresponding VWF- and platelet-binding data are shown in the Figure 2 of the main article.
- Figure S1. Proposed model of collagen III binding to VWF A3 (PDF, 1.94 MB) -
VWF A3–collagen complexes obtained by docking suggest that residues from all 3 strands of collagen contribute to the VWF-binding process. Shown are the 2 highest-ranking docking solutions. Collagen residues Arg572, Hyp575, Val577, and Phe580 that are essential for VWF binding, according to the Ala scan (see Figure 2 of the main article), are shown in stick representation. Residues located in different collagen chains are colored differently (magenta, red, or green). Selected interacting residues of VWF A3 are depicted as blue sticks. A hydrogen bond between the His1023 side chain and a main chain carbonyl of collagen is indicated by a dotted line. The VWF A3 domain is depicted in light-gray ribbon representation; the collagen triple helix, as dark gray tubes. The complex structures were obtained by docking (using HADDOCK version 1.31 10/7-conformation models their side chains optimized using CNS software2 of residues 564 to 587 of human collagen III) the sequence GPOGPSGPRGQOGVMGFOGPKGND onto 2 crystal structures: 1AO33 and 1ATZ4 of the VWF A3 domain. Both models suggest the following interactions to be important: R572 and O575 of one collagen strand interact with A3 D979 (salt bridge) and D979 or I978 (hydrogen bond), respectively; a second collagen strand, R572, with A3 E1001 (salt bridge); and the third collagen strand, V577 and F580, with a hydrophobic patch on A3 composed of I978, F1013, and Y1017.
REFERENCES
1. Dominguez C, Boelens R, Bonvin AM. HADDOCK: a protein-protein docking approach based on biochemical or biophysical information. J Am Chem Soc. 2003;125:1731-1737.
2. Brunger AT, Adams PD, Clore GM, et al. Crystallography & NMR system: a new software suite for macromolecular structure determination. Acta Crystallogr D Biol Crystallogr. 1998;54(pt 5):905-921.
3. Bienkowska J, Cruz M, Atiemo A, Handin R, Liddington R. The von Willebrand factor A3 domain does not contain a metal ion-dependent adhesion site motif. J Biol Chem. 1997;272:25162-25167.
4. Huizinga EG, van der Plas RM, Kroon J, Sixma JJ, Gros P. Crystal structure of the A3 domain of human von Willebrand factor: implications for collagen binding. Structure. 1997;5:1147-1156.
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