Comparison of the Wild-Type α-Amylase and Its Variant Enzymes in Bacillus amyloliquefaciens in Activity and Thermal Stability, and Insights into Engineering the Thermal Stability of Bacillus α-Amylase
| Autor: | Hiroshi Oneda, Kuniyo Inouye, Yoshiki Mouri, Masashi Minoda, Seunjae Lee |
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| Rok vydání: | 2006 |
| Předmět: |
DNA
Bacterial Bacillus amyloliquefaciens Stereochemistry Molecular Sequence Data Bacillus Protein Engineering Models Biological Biochemistry Calcium Chloride Ion binding Enzyme Stability Thermal stability Amino Acid Sequence Enzyme kinetics Amylase Molecular Biology chemistry.chemical_classification Aspartic Acid Base Sequence biology Chemistry Temperature Sequence Analysis DNA General Medicine biology.organism_classification Enzyme Activation Dissociation constant Enzyme Mutation biology.protein Calcium alpha-Amylases Alpha-amylase |
| Zdroj: | The Journal of Biochemistry. 139:1007-1015 |
| ISSN: | 1756-2651 0021-924X |
| DOI: | 10.1093/jb/mvj107 |
| Popis: | The starch hydrolysis activity and thermal stability of Bacillus amyloliquefaciens alpha-amylase (wild-type enzyme or WT) and its variant enzymes, designated as M77, M111, and 21B, were compared. All have an optimal pH at around 6, as well as almost the same reaction rates and Km and kcat values. The optimal temperature in the absence of Ca2+ ions is 60 degrees C for WT and M77 and 40 degrees C for M111 and 21B. Those of M111 and 21B rose to 50-60 degrees C upon the addition of 5 mM CaCl2, while those of WT and M77 did not change. The dissociation constants Kd for Ca2+ to WT and M77 are much lower than those of M111 and 21B. Asp233 in WT is replaced by Asn in M111 and 21B, while it is retained in M77, suggesting that Asp233 is involved in the thermal stability of the enzyme through Ca2+ ion binding. These findings provide insight into engineering the thermal stability of B. amyloliquefaciens alpha-amylase, which would be useful for its applications in the baking industry and in glucose manufacturing. |
| Databáze: | OpenAIRE |
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