Protein thermostability above 100°C: A key role for ionic interactions
| Autor: | David W. Rice, Frank T. Robb, Costantino Vetriani, Nicola Tolliday, Dennis L. Maeder, Kitty S. P. Yip, Timothy J. Stillman, K. Linda Britton, Horst H. Klump |
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| Rok vydání: | 1998 |
| Předmět: |
Models
Molecular Hot Temperature Molecular Sequence Data Crystallography X-Ray Homology (biology) Glutamate Dehydrogenase Enzyme Stability Amino Acid Sequence Thermococcus litoralis Peptide sequence DNA Primers Thermostability Ions chemistry.chemical_classification Multidisciplinary Base Sequence Calorimetry Differential Scanning Sequence Homology Amino Acid biology Glutamate dehydrogenase Biological Sciences biology.organism_classification Hyperthermophile Enzyme Biochemistry chemistry Mutagenesis Site-Directed Pyrococcus furiosus |
| Zdroj: | Proceedings of the National Academy of Sciences. 95:12300-12305 |
| ISSN: | 1091-6490 0027-8424 |
| DOI: | 10.1073/pnas.95.21.12300 |
| Popis: | The discovery of hyperthermophilic microorganisms and the analysis of hyperthermostable enzymes has established the fact that multisubunit enzymes can survive for prolonged periods at temperatures above 100°C. We have carried out homology-based modeling and direct structure comparison on the hexameric glutamate dehydrogenases from the hyperthermophiles Pyrococcus furiosus and Thermococcus litoralis whose optimal growth temperatures are 100°C and 88°C, respectively, to determine key stabilizing features. These enzymes, which are 87% homologous, differ 16-fold in thermal stability at 104°C. We observed that an intersubunit ion-pair network was substantially reduced in the less stable enzyme from T. litoralis , and two residues were then altered to restore these interactions. The single mutations both had adverse effects on the thermostability of the protein. However, with both mutations in place, we observed a fourfold improvement of stability at 104°C over the wild-type enzyme. The catalytic properties of the enzymes were unaffected by the mutations. These results suggest that extensive ion-pair networks may provide a general strategy for manipulating enzyme thermostability of multisubunit enzymes. However, this study emphasizes the importance of the exact local environment of a residue in determining its effects on stability. |
| Databáze: | OpenAIRE |
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