Role of Arginine 129 in Heparin Binding and Activation of Antithrombin
| Autor: | Steven T. Olson, Umesh R. Desai, Ingemar Björk, Susan Clark Bock, Richard Swanson |
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| Rok vydání: | 2000 |
| Předmět: |
Models
Molecular Heparin Protein Conformation Chemistry Antithrombin Allosteric regulation Thrombin Cell Biology Plasma protein binding Serpin Arginine Biochemistry Antithrombins Recombinant Proteins carbohydrates (lipids) Kinetics Protein structure medicine Molecular Biology Protein Binding medicine.drug |
| Zdroj: | Journal of Biological Chemistry. 275:18976-18984 |
| ISSN: | 0021-9258 |
| Popis: | The contribution of Arg(129) of the serpin, antithrombin, to the mechanism of allosteric activation of the protein by heparin was determined from the effect of mutating this residue to either His or Gln. R129H and R129Q antithrombins bound pentasaccharide and full-length heparins containing the antithrombin recognition sequence with similar large reductions in affinity ranging from 400- to 2500-fold relative to the control serpin, corresponding to a loss of 28-35% of the binding free energy. The salt dependence of pentasaccharide binding showed that the binding defect of the mutant serpin resulted from the loss of approximately 2 ionic interactions, suggesting that Arg(129) binds the pentasaccharide cooperatively with other residues. Rapid kinetic studies showed that the mutation minimally affected the initial low affinity binding of heparin to antithrombin, but greatly affected the subsequent conformational activation of the serpin leading to high affinity heparin binding, although not enough to disfavor activation. Consistent with these findings, the mutant antithrombin was normally activated by heparin for accelerated inhibition of factor Xa and thrombin. These results support an important role for Arg(129) in an induced-fit mechanism of heparin activation of antithrombin wherein conformational activation of the serpin positions Arg(129) and other residues for cooperative interactions with the heparin pentasaccharide so as to lock the serpin in the activated state. |
| Databáze: | OpenAIRE |
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