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Structural determinants in the stability of the serpin/proteinase complex
B De Taeye1, K Verbeke, G Compernolle
1Laboratory for Pharmaceutical Biology and Phytopharmacology, Faculty of Pharmaceutical Sciences, Katholieke Universiteit Leuven, E Van Evenstraat 4, Leuven BE-3000, Belgium.
Biochemical and Biophysical Research Communications
|August 2, 2003
Summary
Neutralizing antibodies targeting specific sites on plasminogen activator inhibitor-1 (PAI-1) can alter the stability of PAI-1/proteinase complexes. This suggests antibody interference with proteinase docking sites influences complex stability.
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Serpins, like PAI-1, regulate serine proteinases via stable complex formation.
- Understanding PAI-1/proteinase complex dynamics is crucial for protease inhibition mechanisms.
Purpose of the Study:
- To investigate how PAI-1 neutralizing antibodies (MA) affect the stability of PAI-1/proteinase complexes.
- To determine the impact of antibody binding site on complex stability.
Main Methods:
- Utilized PAI-1 variants with destabilized reactive center loops.
- Employed specific PAI-1 neutralizing antibodies (MA-8H9D4, MA-33H1F7, MA-55F4C12) targeting different regions.
- Assessed the stability of PAI-1/proteinase complexes (t-PA, u-PA) in the presence of antibodies.
Main Results:
- MA-8H9D4 showed no effect on complex stability.
- MA-33H1F7 significantly increased PAI-1/t-PA (7-fold) and PAI-1/u-PA (3-fold) complex stability.
- MA-55F4C12 stabilized PAI-1/t-PA (3-fold) but destabilized PAI-1/u-PA (2-fold) complexes.
Conclusions:
- Antibody interference with the proteinase docking site in preformed serpin/proteinase complexes can modulate intrinsic stability.
- Hypothesized that altered torsion of the reactive center loop (RCL) on the proteinase catalytic triad underlies these stability changes.