Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose
Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequen...
Ausführliche Beschreibung
Gespeichert in:
| Autor*in: |
Kashima, Yasuhiro [verfasserIn] Mori, Kazushige [verfasserIn] Fukada, Harumi [verfasserIn] Ishikawa, Kazuhiko [verfasserIn] |
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| Format: |
E-Artikel |
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| Sprache: |
Englisch |
| Erschienen: |
2004 |
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| Schlagwörter: |
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| Übergeordnetes Werk: |
Enthalten in: Extremophiles - Springer-Verlag, 2001, 9(2004), 1 vom: 16. Sept., Seite 37-43 |
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| Übergeordnetes Werk: |
volume:9 ; year:2004 ; number:1 ; day:16 ; month:09 ; pages:37-43 |
| Links: |
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| DOI / URN: |
10.1007/s00792-004-0418-z |
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SPR007854307 |
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| 520 | |a Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. | ||
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10.1007/s00792-004-0418-z doi (DE-627)SPR007854307 (SPR)s00792-004-0418-z-e DE-627 ger DE-627 rakwb eng Kashima, Yasuhiro verfasserin aut Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose 2004 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. Active site (dpeaa)DE-He213 Cellulase (dpeaa)DE-He213 Chitinase (dpeaa)DE-He213 Disulfide bond (dpeaa)DE-He213 Endoglucanase (dpeaa)DE-He213 Hyperthermophile (dpeaa)DE-He213 Site-directed mutagenesis (dpeaa)DE-He213 Mori, Kazushige verfasserin aut Fukada, Harumi verfasserin aut Ishikawa, Kazuhiko verfasserin aut Enthalten in Extremophiles Springer-Verlag, 2001 9(2004), 1 vom: 16. Sept., Seite 37-43 (DE-627)SPR007852657 nnns volume:9 year:2004 number:1 day:16 month:09 pages:37-43 https://dx.doi.org/10.1007/s00792-004-0418-z lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER AR 9 2004 1 16 09 37-43 |
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10.1007/s00792-004-0418-z doi (DE-627)SPR007854307 (SPR)s00792-004-0418-z-e DE-627 ger DE-627 rakwb eng Kashima, Yasuhiro verfasserin aut Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose 2004 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. Active site (dpeaa)DE-He213 Cellulase (dpeaa)DE-He213 Chitinase (dpeaa)DE-He213 Disulfide bond (dpeaa)DE-He213 Endoglucanase (dpeaa)DE-He213 Hyperthermophile (dpeaa)DE-He213 Site-directed mutagenesis (dpeaa)DE-He213 Mori, Kazushige verfasserin aut Fukada, Harumi verfasserin aut Ishikawa, Kazuhiko verfasserin aut Enthalten in Extremophiles Springer-Verlag, 2001 9(2004), 1 vom: 16. Sept., Seite 37-43 (DE-627)SPR007852657 nnns volume:9 year:2004 number:1 day:16 month:09 pages:37-43 https://dx.doi.org/10.1007/s00792-004-0418-z lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER AR 9 2004 1 16 09 37-43 |
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10.1007/s00792-004-0418-z doi (DE-627)SPR007854307 (SPR)s00792-004-0418-z-e DE-627 ger DE-627 rakwb eng Kashima, Yasuhiro verfasserin aut Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose 2004 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. Active site (dpeaa)DE-He213 Cellulase (dpeaa)DE-He213 Chitinase (dpeaa)DE-He213 Disulfide bond (dpeaa)DE-He213 Endoglucanase (dpeaa)DE-He213 Hyperthermophile (dpeaa)DE-He213 Site-directed mutagenesis (dpeaa)DE-He213 Mori, Kazushige verfasserin aut Fukada, Harumi verfasserin aut Ishikawa, Kazuhiko verfasserin aut Enthalten in Extremophiles Springer-Verlag, 2001 9(2004), 1 vom: 16. Sept., Seite 37-43 (DE-627)SPR007852657 nnns volume:9 year:2004 number:1 day:16 month:09 pages:37-43 https://dx.doi.org/10.1007/s00792-004-0418-z lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER AR 9 2004 1 16 09 37-43 |
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10.1007/s00792-004-0418-z doi (DE-627)SPR007854307 (SPR)s00792-004-0418-z-e DE-627 ger DE-627 rakwb eng Kashima, Yasuhiro verfasserin aut Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose 2004 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. Active site (dpeaa)DE-He213 Cellulase (dpeaa)DE-He213 Chitinase (dpeaa)DE-He213 Disulfide bond (dpeaa)DE-He213 Endoglucanase (dpeaa)DE-He213 Hyperthermophile (dpeaa)DE-He213 Site-directed mutagenesis (dpeaa)DE-He213 Mori, Kazushige verfasserin aut Fukada, Harumi verfasserin aut Ishikawa, Kazuhiko verfasserin aut Enthalten in Extremophiles Springer-Verlag, 2001 9(2004), 1 vom: 16. Sept., Seite 37-43 (DE-627)SPR007852657 nnns volume:9 year:2004 number:1 day:16 month:09 pages:37-43 https://dx.doi.org/10.1007/s00792-004-0418-z lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER AR 9 2004 1 16 09 37-43 |
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10.1007/s00792-004-0418-z doi (DE-627)SPR007854307 (SPR)s00792-004-0418-z-e DE-627 ger DE-627 rakwb eng Kashima, Yasuhiro verfasserin aut Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose 2004 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. Active site (dpeaa)DE-He213 Cellulase (dpeaa)DE-He213 Chitinase (dpeaa)DE-He213 Disulfide bond (dpeaa)DE-He213 Endoglucanase (dpeaa)DE-He213 Hyperthermophile (dpeaa)DE-He213 Site-directed mutagenesis (dpeaa)DE-He213 Mori, Kazushige verfasserin aut Fukada, Harumi verfasserin aut Ishikawa, Kazuhiko verfasserin aut Enthalten in Extremophiles Springer-Verlag, 2001 9(2004), 1 vom: 16. Sept., Seite 37-43 (DE-627)SPR007852657 nnns volume:9 year:2004 number:1 day:16 month:09 pages:37-43 https://dx.doi.org/10.1007/s00792-004-0418-z lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER AR 9 2004 1 16 09 37-43 |
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Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose Active site (dpeaa)DE-He213 Cellulase (dpeaa)DE-He213 Chitinase (dpeaa)DE-He213 Disulfide bond (dpeaa)DE-He213 Endoglucanase (dpeaa)DE-He213 Hyperthermophile (dpeaa)DE-He213 Site-directed mutagenesis (dpeaa)DE-He213 |
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misc Active site misc Cellulase misc Chitinase misc Disulfide bond misc Endoglucanase misc Hyperthermophile misc Site-directed mutagenesis |
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misc Active site misc Cellulase misc Chitinase misc Disulfide bond misc Endoglucanase misc Hyperthermophile misc Site-directed mutagenesis |
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misc Active site misc Cellulase misc Chitinase misc Disulfide bond misc Endoglucanase misc Hyperthermophile misc Site-directed mutagenesis |
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Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose |
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| title_full |
Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose |
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Kashima, Yasuhiro |
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2004 |
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Kashima, Yasuhiro Mori, Kazushige Fukada, Harumi Ishikawa, Kazuhiko |
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analysis of the function of a hyperthermophilic endoglucanase from pyrococcus horikoshii that hydrolyzes crystalline cellulose |
| title_auth |
Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose |
| abstract |
Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. |
| abstractGer |
Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. |
| abstract_unstemmed |
Abstract A hyperthermophilic β-1,4 endoglucanase was identified in Pyrococcus horikoshii, a hyperthermophilic archaeon. In order to clarify the function of the protein in detail, structural and catalytic site studies were performed using protein engineering. By removing some of the C-terminal sequence of the ORF of the endoglucanase (PH1171), two types of recombinant proteins were expressed from one ORF, using Escherichia coli. One exhibited endoglucanase activity, and the other did not. An SD-like sequence was identified in the ORF of the endoglucanase. By removing the SD-like sequence without changing the amino acid sequence of the endoglucanase, one recombinant endoglucanase was prepared effectively from E. coli. From the analysis of the N- and C-terminal regions of the ORF, this endoglucanase appears to be a secreted and membrane-binding enzyme of P. horikoshii. A mutation analysis of the endoglucanase, using the synthetic substrate, indicated that Glu342 is a candidate for the active center and plays a critical role in the activity of the enzyme. Additional catalytic amino acid residues were not found. These results indicate that the catalytic residue of the enzyme is different from that of typical family 5 endoglucanase, even though it has a high homology to the endoglucanase from Acidothermus celluloliticus. The activity of the enzyme, using carboxy methylcellulose and crystalline cellulose as the substrates, was increased, but not for a synthetic low-molecular substrate when a carbohydrate-binding module of chitinase from P. furiosus was added to the C-terminal region. |
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Analysis of the function of a hyperthermophilic endoglucanase from Pyrococcus horikoshii that hydrolyzes crystalline cellulose |
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