Mutagenesis of the bovSERPINA3-3 demonstrates the requirement of aspartate-371 for intermolecular interaction and formation of dimers
| Autor: | Laure Bremaud, Christophe Combet, A Péré-Brissaud, Nathalie Duprat, E Pinault, Ahmed Ouali, Patrick Pélissier, Xavier Blanchet, Didier Delourme, Abderrahman Maftah |
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| Rok vydání: | 2012 |
| Předmět: |
0303 health sciences
animal structures Intermolecular force Serpin Biology medicine.disease_cause Biochemistry 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine chemistry Polymerization Aspartic acid medicine Biophysics Sodium dodecyl sulfate Molecular Biology Escherichia coli Reactive center Polyacrylamide gel electrophoresis 030217 neurology & neurosurgery 030304 developmental biology |
| Zdroj: | Protein Science. 21:977-986 |
| ISSN: | 0961-8368 |
| DOI: | 10.1002/pro.2078 |
| Popis: | The family of serpins is known to fold into a metastable state that is required for the proteinase inhibition mechanism. One of the consequences of this conformational flexibility is the tendency of some mutated serpins to form polymers, which occur through the insertion of the reactive center loop of one serpin molecule into the A-sheet of another. This “A-sheet polymerization” has remained an attractive explanation for the molecular mechanism of serpinopathies. Polymerization of serpins can also take place in vitro under certain conditions (e.g., pH or temperature). Surprisingly, on sodium dodecyl sulfate/polyacrylamide gel electrophoresis, bovSERPINA3-3 extracted from skeletal muscle or expressed in Escherichia coli was mainly observed as a homodimer. Here, in this report, by site-directed mutagenesis of recombinant bovSERPINA3-3, with substitution D371A, we demonstrate the importance of D371 for the intermolecular linkage observed in denaturing and reducing conditions. This residue influences the electrophoretic and conformational properties of bovSERPINA3-3. By structural modeling of mature bovSERPINA3-3, we propose a new “non-A-sheet swap” model of serpin homodimer in which D371 is involved at the molecular interface. |
| Databáze: | OpenAIRE |
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