ArticleApplied and environmental microbiology2003
Directed evolution of Thermus maltogenic amylase toward enhanced thermal resistance.
Article in Applied and environmental microbiology, 2003. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 16 papers.
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Who cites it
16 citing papers in PubMed, 100 citations in OpenAlex.
- Helicobacter pylori cholesteryl α-glucosides contribute to its pathogenicity and immune response by natural killer T cells.PloS one · 2013Trial
- Enhancing stability of enzymes for industrial applications: molecular insights and emerging approaches.World journal of microbiology & biotechnology · 2025Review
- Nature-inspired Enzyme engineering and sustainable catalysis: biochemical clues from the world of plants and extremophiles.Frontiers in bioengineering and biotechnology · 2023Review
- A Review of Advanced Molecular Engineering Approaches to Enhance the Thermostability of Enzyme Breakers: From Prospective of Upstream Oil and Gas Industry.International journal of molecular sciences · 2022Review
- Review: Engineering of thermostable enzymes for industrial applications.APL bioengineering · 2018Review
- Bacterial and Archaeal α-Amylases: Diversity and Amelioration of the Desirable Characteristics for Industrial Applications.Frontiers in microbiology · 2016Review
- Understanding thermostability factors of Aspergillus niger PhyA phytase: a molecular dynamics study.The protein journal · 2013Article
- Association of novel domain in active site of archaic hyperthermophilic maltogenic amylase from Staphylothermus marinus.The Journal of biological chemistry · 2012Article
- Improvement in the thermostability of D-psicose 3-epimerase from Agrobacterium tumefaciens by random and site-directed mutagenesis.Applied and environmental microbiology · 2011Article
- Enzymes in food processing: a condensed overview on strategies for better biocatalysts.Enzyme research · 2010Article
- Structure of Bacillus amyloliquefaciens alpha-amylase at high resolution: implications for thermal stability.Acta crystallographica. Section F, Structural biology and crystallization communications · 2010Article
- Alpha1,4GlcNAc-capped mucin-type O-glycan inhibits cholesterol alpha-glucosyltransferase from Helicobacter pylori and suppresses H. pylori growth.Glycobiology · 2008Article
- Drosophila deoxyribonucleoside kinase mutants with enhanced ability to phosphorylate purine analogs.Gene therapy · 2007Article
- Potential and utilization of thermophiles and thermostable enzymes in biorefining.Microbial cell factories · 2007Article
- Probing the substrate specificity of aminopyrrolnitrin oxygenase (PrnD) by mutational analysis.Journal of bacteriology · 2006Article
- Laboratory-directed protein evolution.Microbiology and molecular biology reviews : MMBR · 2005Review
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Authors and funding
10 authors at 5 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The thermostability of maltogenic amylase from Thermus sp. strain IM6501 (ThMA) was improved greatly by random mutagenesis using DNA shuffling. Four rounds of DNA shuffling and subsequent recombination of the mutations produced the highly thermostable mutant enzyme ThMA-DM, which had a total of seven individual mutations. The seven amino acid substitutions in ThMA-DM were identified as R26Q, S169N, I333V, M375T, A398V, Q411L, and P453L. The optimal reaction temperature of the recombinant enzyme was 75 degrees C, which was 15 degrees C higher than that of wild-type ThMA, and the melting temperature, as determined by differential scanning calorimetry, was increased by 10.9 degrees C. The half-life of ThMA-DM was 172 min at 80 degrees C, a temperature at which wild-type ThMA was completely inactivated in less than 1 min. Six mutations that were generated during the evolutionary process did not significantly affect the specific activity of the enzyme, while the M375T mutation decreased activity to 23% of the wild-type level. The molecular interactions of the seven mutant residues that contributed to the increased thermostability of the mutant enzyme with other adjacent residues were examined by comparing the modeled tertiary structure of ThMA-DM with those of wild-type ThMA and related enzymes. The A398V and Q411L substitutions appeared to stabilize the enzyme by enhancing the interdomain hydrophobic interactions. The R26Q and P453L substitutions led potentially to the formation of genuine hydrogen bonds. M375T, which was located near the active site of ThMA, probably caused a conformational or dynamic change that enhanced thermostability but reduced the specific activity of the enzyme.
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