Single mutations of residues outside the active center of the xylanase Xys1Î fromStreptomyces halstediiJM8 affect its activity
| Autor: | Ramón I. Santamaría, Valery L. Shnyrov, Alberto Ruiz-Arribas, José M. Fernández-Abalos, Javier De Las Rivas, Sonia Rodríguez, Margarita Díaz |
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| Rok vydání: | 2004 |
| Předmět: |
Models
Molecular Protein Conformation DNA Mutational Analysis Molecular Sequence Data Mutant Mutation Missense Mutagenesis (molecular biology technique) Biology Microbiology Streptomyces Active center Bacterial Proteins Enzyme Stability Genetics Point Mutation Amino Acid Sequence Molecular Biology chemistry.chemical_classification Binding Sites Temperature Wild type biology.organism_classification Xylosidases Enzyme Amino Acid Substitution chemistry Biochemistry Mutagenesis Mutation Xylanase Xylans Specific activity Sequence Alignment |
| Zdroj: | FEMS Microbiology Letters. 240:237-243 |
| ISSN: | 1574-6968 0378-1097 |
| DOI: | 10.1016/j.femsle.2004.09.032 |
| Popis: | Mutagenesis of the xylanase Xys1 of Streptomyces halstedii JM8 has been done by error prone PCR. Mutants with modified hydrolytic activity were isolated, the recombinant variant proteins purified and the catalytic activities of each one determined and compared with the wild type enzyme. Two of the isolated single point mutants, m1 (G133D) and m8 (N148D), showed 22-25% increase in specific activity towards xylan compared to wild type xylanase. Two other mutants, m5a (D175A) and m7 (T160A), showed a significant reduction in specific activity of 40-50% with respect to the wild type enzyme. These residues are mainly located in the beta alpha-loops of the xylanase, the region showing the main structural divergences within family 10 of xylanases. This study shows the usefulness of random mutagenesis to point out some key residues not directly involved in the active center, but in which mutation produces subtle structural rearrangements affecting the enzymatic function. |
| Databáze: | OpenAIRE |
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