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Ionic interactions in the formation of the thrombin-hirudin complex
A Betz1, J Hofsteenge, S R Stone
1Friedrich Miescher-Institut, Basel, Switzerland.
The Biochemical Journal
|May 1, 1991
Summary
Site-directed mutagenesis revealed varying contributions of hirudin
Area of Science:
- Biochemistry
- Molecular Biology
- Protein-protein interactions
Background:
- Hirudin is a potent inhibitor of thrombin.
- The C-terminus of hirudin plays a crucial role in thrombin binding.
- Understanding these interactions is key for drug development.
Purpose of the Study:
- To investigate the role of individual acidic C-terminal residues of hirudin in alpha-thrombin complex formation.
- To quantify the binding energy contributions of these residues.
Main Methods:
- Site-directed mutagenesis was employed to alter specific acidic residues in hirudin's C-terminus.
- Binding energy contributions were assessed through biophysical methods.
Main Results:
- The contribution of acidic residues to binding energy ranged from 2.3 to 5.9 kJ/mol.
- Asp-55 replacement had a surprisingly small effect on binding energy.
- Mutational effects were not always additive, particularly for Glu-57 and Glu-58.
Conclusions:
- Acidic C-terminal residues of hirudin contribute differentially to thrombin binding energy.
- The crystal structure predictions of residue importance were not fully supported by experimental data.
- Non-additive effects in multiple mutations highlight complex interaction dynamics.