Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph
Cross-linked enzyme aggregates (CLEAs) of the Y509E mutant of glycoside hydrolase family 52 β-xylosidase from Geobacillus stearothermophilus with dual activity of β-xylosidase and xylanase (XynB2Y509E) were prepared. Ammonium sulfate was used as the precipitant agent, and glutaraldehyde as cross-lin...
| Main Authors: | , , , , , , |
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| Format: | info:eu-repo/semantics/article |
| Language: | English |
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2021
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| Subjects: | |
| Online Access: | http://hdl.handle.net/10835/9312 |
| _version_ | 1789406736387407872 |
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| author | Romero, Gabriela Contreras Moyeja, Lellys Mariela Aguirre, Carolina Wilkesman, Jeff Clemente Jiménez, María José Rodríguez Vico, Felipe Las Heras Vázquez, Francisco Javier de |
| author_facet | Romero, Gabriela Contreras Moyeja, Lellys Mariela Aguirre, Carolina Wilkesman, Jeff Clemente Jiménez, María José Rodríguez Vico, Felipe Las Heras Vázquez, Francisco Javier de |
| author_sort | Romero, Gabriela |
| collection | DSpace |
| description | Cross-linked enzyme aggregates (CLEAs) of the Y509E mutant of glycoside hydrolase family 52 β-xylosidase from Geobacillus stearothermophilus with dual activity of β-xylosidase and xylanase (XynB2Y509E) were prepared. Ammonium sulfate was used as the precipitant agent, and glutaraldehyde as cross-linking agent. The optimum conditions were found to be 90% ammonium sulfate, 12.5 mM glutaraldehyde, 3 h of cross-linking reaction at 25 °C, and pH 8.5. Under these (most effective) conditions, XynB2Y509E-CLEAs retained 92.3% of their original β-xylosidase activity. Biochemical characterization of both crude and immobilized enzymes demonstrated that the maximum pH and temperature after immobilization remained unchanged (pH 6.5 and 65 °C). Moreover, an improvement in pH stability and thermostability was also found after immobilization. Analysis of kinetic parameters shows that the Km value of XynB2Y509E-CLEAs obtained was slightly higher than that of free XynB2Y509E (1.2 versus 0.9 mM). Interestingly, the xylanase activity developed by the mutation was also conserved after the immobilization process. |
| format | info:eu-repo/semantics/article |
| id | oai:repositorio.ual.es:10835-9312 |
| institution | Universidad de Cuenca |
| language | English |
| publishDate | 2021 |
| record_format | dspace |
| spelling | oai:repositorio.ual.es:10835-93122023-04-12T19:48:25Z Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph Romero, Gabriela Contreras Moyeja, Lellys Mariela Aguirre, Carolina Wilkesman, Jeff Clemente Jiménez, María José Rodríguez Vico, Felipe Las Heras Vázquez, Francisco Javier de β-xylosidase thermostability CLEAs G. stearothermophilus xylanase Cross-linked enzyme aggregates (CLEAs) of the Y509E mutant of glycoside hydrolase family 52 β-xylosidase from Geobacillus stearothermophilus with dual activity of β-xylosidase and xylanase (XynB2Y509E) were prepared. Ammonium sulfate was used as the precipitant agent, and glutaraldehyde as cross-linking agent. The optimum conditions were found to be 90% ammonium sulfate, 12.5 mM glutaraldehyde, 3 h of cross-linking reaction at 25 °C, and pH 8.5. Under these (most effective) conditions, XynB2Y509E-CLEAs retained 92.3% of their original β-xylosidase activity. Biochemical characterization of both crude and immobilized enzymes demonstrated that the maximum pH and temperature after immobilization remained unchanged (pH 6.5 and 65 °C). Moreover, an improvement in pH stability and thermostability was also found after immobilization. Analysis of kinetic parameters shows that the Km value of XynB2Y509E-CLEAs obtained was slightly higher than that of free XynB2Y509E (1.2 versus 0.9 mM). Interestingly, the xylanase activity developed by the mutation was also conserved after the immobilization process. 2021-01-18T09:31:20Z 2021-01-18T09:31:20Z 2021-01-16 info:eu-repo/semantics/article 1420-3049 http://hdl.handle.net/10835/9312 en https://www.mdpi.com/1420-3049/26/2/451 Attribution-NonCommercial-NoDerivatives 4.0 Internacional http://creativecommons.org/licenses/by-nc-nd/4.0/ info:eu-repo/semantics/openAccess |
| spellingShingle | β-xylosidase thermostability CLEAs G. stearothermophilus xylanase Romero, Gabriela Contreras Moyeja, Lellys Mariela Aguirre, Carolina Wilkesman, Jeff Clemente Jiménez, María José Rodríguez Vico, Felipe Las Heras Vázquez, Francisco Javier de Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph |
| title | Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph |
| title_full | Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph |
| title_fullStr | Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph |
| title_full_unstemmed | Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph |
| title_short | Characterization of Cross-Linked Enzyme Aggregates of the Y509E Mutant of a Glycoside Hydrolase Family 52 β-xylosidase from G. stearothermoph |
| title_sort | characterization of cross-linked enzyme aggregates of the y509e mutant of a glycoside hydrolase family 52 β-xylosidase from g. stearothermoph |
| topic | β-xylosidase thermostability CLEAs G. stearothermophilus xylanase |
| url | http://hdl.handle.net/10835/9312 |
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