| DOI | Resolve DOI: https://doi.org/10.1128/AEM.07576-11 |
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| Author | Search for: Yang, T.C.1; Search for: Legault, S.1; Search for: Kayiranga, E.A.; Search for: Kumaran, J.1; Search for: Ishikawa, K.; Search for: Sung, W.L.1 |
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| Affiliation | - National Research Council of Canada. NRC Institute for Biological Sciences
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| Format | Text, Article |
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| Subject | Active site; Bioinformatics analysis; C-terminal sequences; Core structure; Crystalline cellulose; Glucosidase; High temperature; Hydrolyzing activity; Hyperthermophilic archaeon; Hyperthermophilic endoglucanase; Hyperthermostability; Key residues; N-terminals; PH stability; Pyrococcus horikoshii; TIM barrels; Triosephosphate isomerase; Wild types; Bioinformatics; Cellulose; Enzymes; Ethanol; Glucose; Hydrolysis; Industrial applications; Crystal structure; cellulase; bioinformatics; cellulose; enzyme activity; gene expression; high temperature; thermophilic bacterium; amino acid sequence; chemistry; enzyme stability; enzymology; gene deletion; genetics; heat; molecular genetics; pH; protein conformation; protein stability; protein tertiary structure; Pyrococcus horikoshii; Amino Acid Sequence; Cellulase; Enzyme Stability; Hot Temperature; Hydrogen-Ion Concentration; Molecular Sequence Data; Protein Conformation; Protein Stability; Protein Structure, Tertiary; Pyrococcus horikoshii; Sequence Deletion; Archaea; Pyrococcus horikoshii |
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| Abstract | The β-1,4-endoglucanase (EC 3.2.1.4) from the hyperthermophilic archaeon Pyrococcus horikoshii (EGPh) has strong hydrolyzing activity toward crystalline cellulose. When EGPh is used in combination with β-glucosidase (EC 3.2.1.21), cellulose is completely hydrolyzed to glucose at high temperature, suggesting great potential for EGPh in bioethanol industrial applications. The crystal structure of EGPh shows a triosephosphate isomerase (TIM) (β/α)8-barrel fold with an N-terminal antiparallel β-sheet at the opposite side of the active site and a very short C-terminal sequence outside of the barrel structure. We describe here the function of the peripheral sequences outside of the TIM barrel core structure. Sequential deletions were performed from both N and C termini. The activity, thermostability, and pH stability of the expressed mutants were assessed and compared to the wildtype EGPh enzyme. Our results demonstrate that the TIM barrel core is essential for enzyme activity and that the N-terminal β-sheet is critical for enzyme thermostability. Bioinformatics analyses identified potential key residues which may contribute to enzyme hyperthermostability. © 2012, American Society for Microbiology. |
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| Publication date | 2012 |
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| In | |
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| Language | English |
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| Peer reviewed | Yes |
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| NPARC number | 21269354 |
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| Export citation | Export as RIS |
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| Report a correction | Report a correction (opens in a new tab) |
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| Record identifier | 8840a4f5-f407-4b8f-83ff-c35f4d400135 |
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| Record created | 2013-12-12 |
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| Record modified | 2020-04-21 |
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