Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of Casein
This study aimed to evaluate the performance of a commercial protease (Novo-Pro D (NPD)), both in soluble and immobilized forms, in the hydrolysis of proteins (using casein as model protein). Immobilization of the protease NPD on 6% agarose activated with glyoxyl groups for 24 h at 20 °C and pH 10.0...
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MDPI AG
2020-04-01
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| Series: | Catalysts |
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| Online Access: | https://www.mdpi.com/2073-4344/10/5/466 |
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| author | Laiane Antunes Lopes Paula Kern Novelli Roberto Fernandez-Lafuente Paulo Waldir Tardioli Raquel Lima Camargo Giordano |
| author_facet | Laiane Antunes Lopes Paula Kern Novelli Roberto Fernandez-Lafuente Paulo Waldir Tardioli Raquel Lima Camargo Giordano |
| author_sort | Laiane Antunes Lopes |
| collection | DOAJ |
| description | This study aimed to evaluate the performance of a commercial protease (Novo-Pro D (NPD)), both in soluble and immobilized forms, in the hydrolysis of proteins (using casein as model protein). Immobilization of the protease NPD on 6% agarose activated with glyoxyl groups for 24 h at 20 °C and pH 10.0 allowed preparing immobilized biocatalyst with around 90% immobilization yield, 92% recovered activity versus small substrate, and a thermal stability 5.3-fold higher than the dialyzed soluble enzyme at 50 °C and pH 8.0. Immobilization times longer than 24 h lead to a decrease in the recovered activity and did not improve the biocatalyst stability. At 50 °C and pH 6.5, the immobilized NPD was around 20-fold more stable than the dialyzed soluble protease. Versus casein, the immobilized NDP presented a 10% level of activity, but it allowed hydrolyzing casein (26 g/L) at 50 °C and pH 6.5 up to a 40% degree of hydrolysis (DH) after 2 h reaction, while under the same conditions, only a 34% DH was achieved with soluble NPD. In addition, the immobilized NPD showed good reusability, maintaining the DH of casein for at least ten 2h-reaction batches. |
| first_indexed | 2024-03-10T20:14:52Z |
| format | Article |
| id | doaj.art-da6f9edbc7444e46b224d6eaaf089750 |
| institution | Directory Open Access Journal |
| issn | 2073-4344 |
| language | English |
| last_indexed | 2024-03-10T20:14:52Z |
| publishDate | 2020-04-01 |
| publisher | MDPI AG |
| record_format | Article |
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| spelling | doaj.art-da6f9edbc7444e46b224d6eaaf0897502023-11-19T22:40:38ZengMDPI AGCatalysts2073-43442020-04-0110546610.3390/catal10050466Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of CaseinLaiane Antunes Lopes0Paula Kern Novelli1Roberto Fernandez-Lafuente2Paulo Waldir Tardioli3Raquel Lima Camargo Giordano4Postgraduate Program of Chemical Engineering, Department of Chemical Engineering, Federal University of São Carlos (UFSCar), Rod. Washington Luís, km 235, São Carlos 13565-905, SP, BrazilFaculty of Food Engineering, University of Campinas (UNICAMP), Rua Monteiro Lobato, No. 80, Campinas 13083-862, SP, BrazilDepartamento de Biocatálisis, ICP-CSIC, Campus UAM-CSIC, 28040 Madrid, SpainPostgraduate Program of Chemical Engineering, Department of Chemical Engineering, Federal University of São Carlos (UFSCar), Rod. Washington Luís, km 235, São Carlos 13565-905, SP, BrazilPostgraduate Program of Chemical Engineering, Department of Chemical Engineering, Federal University of São Carlos (UFSCar), Rod. Washington Luís, km 235, São Carlos 13565-905, SP, BrazilThis study aimed to evaluate the performance of a commercial protease (Novo-Pro D (NPD)), both in soluble and immobilized forms, in the hydrolysis of proteins (using casein as model protein). Immobilization of the protease NPD on 6% agarose activated with glyoxyl groups for 24 h at 20 °C and pH 10.0 allowed preparing immobilized biocatalyst with around 90% immobilization yield, 92% recovered activity versus small substrate, and a thermal stability 5.3-fold higher than the dialyzed soluble enzyme at 50 °C and pH 8.0. Immobilization times longer than 24 h lead to a decrease in the recovered activity and did not improve the biocatalyst stability. At 50 °C and pH 6.5, the immobilized NPD was around 20-fold more stable than the dialyzed soluble protease. Versus casein, the immobilized NDP presented a 10% level of activity, but it allowed hydrolyzing casein (26 g/L) at 50 °C and pH 6.5 up to a 40% degree of hydrolysis (DH) after 2 h reaction, while under the same conditions, only a 34% DH was achieved with soluble NPD. In addition, the immobilized NPD showed good reusability, maintaining the DH of casein for at least ten 2h-reaction batches.https://www.mdpi.com/2073-4344/10/5/466enzyme immobilizationenzyme stabilizationproteaseprotein hydrolysisagarose-glyoxyl |
| spellingShingle | Laiane Antunes Lopes Paula Kern Novelli Roberto Fernandez-Lafuente Paulo Waldir Tardioli Raquel Lima Camargo Giordano Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of Casein Catalysts enzyme immobilization enzyme stabilization protease protein hydrolysis agarose-glyoxyl |
| title | Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of Casein |
| title_full | Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of Casein |
| title_fullStr | Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of Casein |
| title_full_unstemmed | Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of Casein |
| title_short | Glyoxyl-Activated Agarose as Support for Covalently Link Novo-Pro D: Biocatalysts Performance in the Hydrolysis of Casein |
| title_sort | glyoxyl activated agarose as support for covalently link novo pro d biocatalysts performance in the hydrolysis of casein |
| topic | enzyme immobilization enzyme stabilization protease protein hydrolysis agarose-glyoxyl |
| url | https://www.mdpi.com/2073-4344/10/5/466 |
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