Publication: Disease-causing mutations in the serpin antithrombin reveal a keydomaincritical for inhibiting protease activities
Authors
Águila Martínez, Sonia ; Izaguirre, G. ; Vicente García, Vicente ; Martínez-Martínez, I. ; Olson, S. T. ; Corral de la Calle, Javier
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Publisher
Elsevier, American Society for Biochemistry and Molecular Biology
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DOI
https://doi.org/10.1074/jbc.M117.787325
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info:eu-repo/semantics/article
Description
Abstract
Antithrombin mainly inhibits factor Xa and thrombin. The reactive center loop (RCL) is crucial for its interactions with its protease targets and is fully inserted into the A-sheet after its cleavage, causing translocation of the covalently linked protease to the opposite end of the A-sheet. Antithrombin variants with altered RCL hinge residues behave as substrates rather than inhibitors, resulting in stoichiometries of inhibition greater than one. Other antithrombin residues have been suggested to interfere with RCL insertion or the stability of the antithrombin–protease complex, but available crystal structures or mutagenesis studies have failed to identify such residues. Here, we characterized two mutations, S365L and I207T, present in individuals with type II antithrombin deficiency and identified a new antithrombin functional domain. S365L did not form stable complexes with thrombin or factor Xa, and the I207T/I207A variants inhibited both proteases with elevated stoichiometries of inhibition. Close proximity of Ile-207 and Ser-365 to the inserted RCL suggested that the preferred reaction of these mutants as protease substrates reflects an effect on the rate of the RCL insertion and protease translocation. However, both residues lie within the final docking site for the protease in the antithrombin–protease complex, supporting the idea that the enhanced substrate reactions may result from an increased dissociation of the final complexes. Our findings demonstrate that the distal end of the antithrombin A-sheet is crucial for the last steps of protease inhibition either by affecting the rate of RCL insertion or through critical interactions with proteases at the end of the A-sheet.
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Citation
Journal of Biological Chemistry, Volume 292, Issue 40(2017),16521-16522.
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