Improved Covalent Immobilization of Horseradish Peroxidase on Macroporous Glycidyl Methacrylate-Based Copolymers
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2012
Authors
Prodanović, OliveraProkopijević, Miloš

Spasojević, Dragica
Stojanović, Željko
Radotić, Ksenija

Knezevic-Jugovic, Zorica D.
Prodanović, Radivoje

Article (Published version)

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A macroporous copolymer of glycidyl methacrylate and ethylene glycol dimethacrylate, poly(GMA-co-EGDMA), with various surface characteristics and mean pore size diameters ranging from 44 to 200 nm was synthesized, modified with 1,2-diaminoethane, and tested as a carrier for immobilization of horseradish peroxidase (HRP) by two covalent methods, glutaraldehyde and periodate. The highest specific activity of around 35 U g(-1) dry weight of carrier was achieved on poly(GMA-co-EGDMA) copolymers with mean pore diameters of 200 and 120 nm by the periodate method. A study of deactivation kinetics at 65 A degrees C and in 80 % dioxane revealed that periodate immobilization also produced an appreciable stabilization of the biocatalyst, while stabilization factor depended strongly on the surface characteristics of the copolymers. HRP immobilized on copolymer with a mean pore diameter of 120 nm by periodate method showing not only the highest specific activity but also good stability was further ...characterized. It appeared that the immobilization resulted in the stabilization of enzyme over a broader pH range while the Michaelis constant value (K (m)) of the immobilized HRP was 10.8 mM, approximately 5.6 times higher than that of the free enzyme. After 6 cycles of repeated use in a batch reactor for pyrogallol oxidation, the immobilized HRP retained 45 % of its original activity.
Keywords:
Macroporous polymers / Copolymerization / Enzymes / Horseradish peroxidase / Covalent immobilization / Morphology / Biological applications of polymersSource:
Applied Biochemistry and Biotechnology, 2012, 168, 5, 1288-1301Publisher:
- Humana Press Inc, Totowa
Funding / projects:
- Study of structure-function relationships in the plant cell wall and modifications of the wall structure by enzyme engineering (RS-173017)
- Allergens, antibodies, enzymes and small physiologically important molecules: design, structure, function and relevance (RS-172049)
DOI: 10.1007/s12010-012-9857-7
ISSN: 0273-2289
PubMed: 22941271
WoS: 000311310300026
Scopus: 2-s2.0-84871904972
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Institution/Community
Hemijski fakultet / Faculty of ChemistryTY - JOUR AU - Prodanović, Olivera AU - Prokopijević, Miloš AU - Spasojević, Dragica AU - Stojanović, Željko AU - Radotić, Ksenija AU - Knezevic-Jugovic, Zorica D. AU - Prodanović, Radivoje PY - 2012 UR - https://cherry.chem.bg.ac.rs/handle/123456789/1552 AB - A macroporous copolymer of glycidyl methacrylate and ethylene glycol dimethacrylate, poly(GMA-co-EGDMA), with various surface characteristics and mean pore size diameters ranging from 44 to 200 nm was synthesized, modified with 1,2-diaminoethane, and tested as a carrier for immobilization of horseradish peroxidase (HRP) by two covalent methods, glutaraldehyde and periodate. The highest specific activity of around 35 U g(-1) dry weight of carrier was achieved on poly(GMA-co-EGDMA) copolymers with mean pore diameters of 200 and 120 nm by the periodate method. A study of deactivation kinetics at 65 A degrees C and in 80 % dioxane revealed that periodate immobilization also produced an appreciable stabilization of the biocatalyst, while stabilization factor depended strongly on the surface characteristics of the copolymers. HRP immobilized on copolymer with a mean pore diameter of 120 nm by periodate method showing not only the highest specific activity but also good stability was further characterized. It appeared that the immobilization resulted in the stabilization of enzyme over a broader pH range while the Michaelis constant value (K (m)) of the immobilized HRP was 10.8 mM, approximately 5.6 times higher than that of the free enzyme. After 6 cycles of repeated use in a batch reactor for pyrogallol oxidation, the immobilized HRP retained 45 % of its original activity. PB - Humana Press Inc, Totowa T2 - Applied Biochemistry and Biotechnology T1 - Improved Covalent Immobilization of Horseradish Peroxidase on Macroporous Glycidyl Methacrylate-Based Copolymers VL - 168 IS - 5 SP - 1288 EP - 1301 DO - 10.1007/s12010-012-9857-7 ER -
@article{ author = "Prodanović, Olivera and Prokopijević, Miloš and Spasojević, Dragica and Stojanović, Željko and Radotić, Ksenija and Knezevic-Jugovic, Zorica D. and Prodanović, Radivoje", year = "2012", abstract = "A macroporous copolymer of glycidyl methacrylate and ethylene glycol dimethacrylate, poly(GMA-co-EGDMA), with various surface characteristics and mean pore size diameters ranging from 44 to 200 nm was synthesized, modified with 1,2-diaminoethane, and tested as a carrier for immobilization of horseradish peroxidase (HRP) by two covalent methods, glutaraldehyde and periodate. The highest specific activity of around 35 U g(-1) dry weight of carrier was achieved on poly(GMA-co-EGDMA) copolymers with mean pore diameters of 200 and 120 nm by the periodate method. A study of deactivation kinetics at 65 A degrees C and in 80 % dioxane revealed that periodate immobilization also produced an appreciable stabilization of the biocatalyst, while stabilization factor depended strongly on the surface characteristics of the copolymers. HRP immobilized on copolymer with a mean pore diameter of 120 nm by periodate method showing not only the highest specific activity but also good stability was further characterized. It appeared that the immobilization resulted in the stabilization of enzyme over a broader pH range while the Michaelis constant value (K (m)) of the immobilized HRP was 10.8 mM, approximately 5.6 times higher than that of the free enzyme. After 6 cycles of repeated use in a batch reactor for pyrogallol oxidation, the immobilized HRP retained 45 % of its original activity.", publisher = "Humana Press Inc, Totowa", journal = "Applied Biochemistry and Biotechnology", title = "Improved Covalent Immobilization of Horseradish Peroxidase on Macroporous Glycidyl Methacrylate-Based Copolymers", volume = "168", number = "5", pages = "1288-1301", doi = "10.1007/s12010-012-9857-7" }
Prodanović, O., Prokopijević, M., Spasojević, D., Stojanović, Ž., Radotić, K., Knezevic-Jugovic, Z. D.,& Prodanović, R.. (2012). Improved Covalent Immobilization of Horseradish Peroxidase on Macroporous Glycidyl Methacrylate-Based Copolymers. in Applied Biochemistry and Biotechnology Humana Press Inc, Totowa., 168(5), 1288-1301. https://doi.org/10.1007/s12010-012-9857-7
Prodanović O, Prokopijević M, Spasojević D, Stojanović Ž, Radotić K, Knezevic-Jugovic ZD, Prodanović R. Improved Covalent Immobilization of Horseradish Peroxidase on Macroporous Glycidyl Methacrylate-Based Copolymers. in Applied Biochemistry and Biotechnology. 2012;168(5):1288-1301. doi:10.1007/s12010-012-9857-7 .
Prodanović, Olivera, Prokopijević, Miloš, Spasojević, Dragica, Stojanović, Željko, Radotić, Ksenija, Knezevic-Jugovic, Zorica D., Prodanović, Radivoje, "Improved Covalent Immobilization of Horseradish Peroxidase on Macroporous Glycidyl Methacrylate-Based Copolymers" in Applied Biochemistry and Biotechnology, 168, no. 5 (2012):1288-1301, https://doi.org/10.1007/s12010-012-9857-7 . .