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Zinc and calcium ions cooperatively modulate ADAMTS13 activity
Patricia J Anderson1, Koichi Kokame, J Evan Sadler
1Howard Hughes Medical Institute, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA.
The Journal of Biological Chemistry
|November 16, 2005
Summary
The metalloproteinase ADAMTS13 activity, crucial for cleaving von Willebrand factor (VWF), is significantly enhanced by zinc and calcium ions. This metal ion dependence highlights key factors influencing VWF multimer processing in plasma.
Area of Science:
- Biochemistry
- Molecular Biology
- Hematology
Background:
- ADAMTS13 is a key metalloproteinase responsible for cleaving von Willebrand factor (VWF) multimers.
- Understanding the regulation of ADAMTS13 activity is crucial for VWF-mediated platelet aggregation and thrombosis.
- Metal ion cofactors are known to influence the activity of many metalloproteinases.
Purpose of the Study:
- To investigate the metal ion dependence of ADAMTS13 activity.
- To characterize the activation of ADAMTS13 by zinc, calcium, and barium ions using both multimeric VWF and a peptide substrate.
- To determine the kinetic parameters of ADAMTS13 cleavage for VWF and a peptide substrate.
Main Methods:
- Assessed ADAMTS13 activity using multimeric VWF and a fluorescent peptide substrate (FRETS-VWF73).
- Examined the effect of various metal ions (Zn2+, Ca2+, Ba2+) on ADAMTS13 activity in plasma and purified systems.
- Determined kinetic parameters (Km, catalytic efficiency) for VWF and FRETS-VWF73 cleavage.
Main Results:
- ADAMTS13 activity was significantly enhanced by zinc (approx. 2-fold in plasma, up to 200-fold with recombinant enzyme) and calcium ions (approx. 3-fold in plasma, up to 200-fold with recombinant enzyme).
- Cooperative activation was observed with both zinc and calcium ions.
- Barium ions stimulated activity in citrated plasma, likely by chelating citrate and releasing bound Ca2+ and Zn2+.
- ADAMTS13 demonstrated a strong preference for VWF over the peptide substrate, suggesting specific structural recognition sites on VWF.
Conclusions:
- Zinc and calcium ions are potent activators of ADAMTS13, with synergistic effects observed.
- The activity of ADAMTS13 is modulated by the presence of chelating agents like citrate in plasma.
- ADAMTS13 exhibits substrate specificity, preferentially cleaving VWF multimers over smaller peptide fragments, indicating recognition of structural features on VWF.