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Published on: August 14, 2017
Identification of ADAMTS13 peptide sequences binding to von Willebrand factor
Takanori Moriki1, Ichiro N Maruyama, Atsuko Igari
1Health Center, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. TakaMoriki@aol.com
Abstract:
ADAMTS13 cleaves multimeric von Willebrand factor (VWF) to regulate VWF-mediated thrombus formation. To search ADAMTS13 peptide sequences binding to VWF, a lambda-phage library expressing various peptides of ADAMTS13 on the surface was screened using VWF either immobilized or in solution under static condition. By the first screening, peptides sharing the C-terminus of spacer domain from Arg(670) to Gln(684) (epitope-A) were selected. To explore additional sites, peptide sequences from the first screening were synthesized and added to the second screening. Consequently, Pro(618) to Glu(641) (epitope-B) in the middle of spacer domain was obtained from immobilized VWF condition. Synthetic epitope-B peptide inhibited the cleavage of VWF by ADAMTS13, while the synthetic epitope-A peptide did not as efficiently as epitope-B. Elimination of four amino acids from either sides of epitope-B terminus markedly reduced the inhibitory effect. These two sites in the spacer domain may play significant roles in binding to VWF.
Insights
Researchers identified two key binding sites on ADAMTS13 (a protein that cleaves von Willebrand factor) that are crucial for regulating blood clot formation. These findings advance understanding of ADAMTS13-VWF interactions.
Area of Science:
- Biochemistry
- Hematology
- Molecular Biology
Background:
- ADAMTS13 enzyme is essential for regulating thrombus formation by cleaving von Willebrand factor (VWF).
- Understanding the specific binding sites of ADAMTS13 on VWF is critical for elucidating its regulatory mechanism.
Purpose of the Study:
- To identify and characterize peptide sequences of ADAMTS13 that bind to VWF.
- To investigate the functional significance of these binding sites in VWF cleavage and regulation of thrombus formation.
Main Methods:
- Screening of a lambda-phage library displaying ADAMTS13 peptides against immobilized and solution-phase VWF.
- Synthesis and functional testing of identified ADAMTS13 peptide sequences (epitope-A and epitope-B).
- Assessing the inhibitory effect of synthetic peptides on VWF cleavage by ADAMTS13.
Main Results:
- Two distinct binding sites within the ADAMTS13 spacer domain were identified: epitope-A (Arg670-Gln684) and epitope-B (Pro618-Glu641).
- Synthetic epitope-B peptide demonstrated significant inhibition of VWF cleavage by ADAMTS13, more so than epitope-A.
- Truncation of epitope-B reduced its inhibitory activity, highlighting the importance of this region.
Conclusions:
- The spacer domain of ADAMTS13 contains critical VWF binding sites, specifically epitope-A and epitope-B.
- Epitope-B plays a significant role in the interaction with VWF and in modulating ADAMTS13 activity.
- These findings provide insights into the molecular mechanisms of ADAMTS13-VWF interaction and thrombus regulation.
