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Probing inhibitor-induced conformational changes along the interface between tissue factor and factor VIIa.
M Osterlund1, R Owenius, K Carlsson
1IFM-Department of Chemistry, Linköping University, Linköping, Sweden.
Biochemistry
|August 2, 2001
Summary
Inhibiting factor VIIa (FVIIa) with active site inhibitors makes the tissue factor (TF) binding interface more rigid. This structural change occurs specifically where FVIIa’s protease domain interacts with TF.
Area of Science:
- Biochemistry
- Molecular Biology
- Hemostasis
Background:
- Activated factor VII (FVIIa) initiates blood clotting by binding to tissue factor (TF).
- Active site-inhibited FVIIa (FVIIai) exhibits higher affinity for TF than FVIIa.
- Understanding the structural dynamics of the FVIIa-TF interaction is crucial for hemostasis research.
Purpose of the Study:
- To compare the interactions of FVIIa and FVIIai with soluble TF (sTF).
- To investigate the structural consequences of active site inhibition on the FVIIa-TF interface.
Main Methods:
- Site-directed mutagenesis of six residues in sTF at the FVIIa binding interface.
- Attachment of fluorescent and spin probes to engineered cysteine residues in sTF.
- Monitoring changes in hydrophobicity, accessibility, and rigidity within the sTF-FVIIa complex using spectroscopic techniques.
Main Results:
- Active site inhibition of FVIIa led to increased rigidity and decreased solvent accessibility in sTF regions interacting with FVIIa's protease domain.
- The FVIIa active site inhibitor induced a less flexible and more compact interface structure.
- The interaction interface between sTF and the light chain of FVIIa remained unaffected by active site occupancy.
Conclusions:
- Active site occupancy in FVIIa significantly alters the structural dynamics of its interface with TF.
- The findings highlight the role of FVIIa's protease domain in mediating TF binding interface rigidity.
- This study provides insights into the allosteric regulation of the FVIIa-TF complex.