Protein‐coated polymer as a matrix for enzyme immobilization: Immobilization of trypsin on bovine serum albumin‐coated allyl glycidyl ether–ethylene glycol dimethacrylate copolymer

Abstract

Allyl glycidyl ether (AGE)-ethylene glycol dimethacrylate (EGDM) copolymer with 25% crosslink density (AGE-25) shows excellent bovine serum albumin (BSA) adsorption (up to 16% (w/w)) at pH 8.0 and the adsorbed BSA is strongly bound. This protein-coated polymer provides a novel matrix with naturally existing functional groups such as thiol, amino, and carboxylic acid that are available for covalent immobilization of functional enzymes. Employing appropriate strategies, trypsin as a model protein was covalently bound to BSA-coated matrix both independently, and in a stepwise manner on the same matrix, with less than 5% loss of enzyme activity during immobilization. Glutaraldehyde crosslinking after immobilization provide stable enzyme preparation with activity of 510 units/g recycled up to six times without loss of enzyme activity. AFM studies reveal that the polymer surface has protein peaks and valleys rather than a uniform monolayer distribution of the protein and the immobilized enzyme preparation can best be described as polymer supported cross-linked enzyme aggregates (CLEAs).

Immobilization of Bovine Serum Albumin onto Porous Poly(vinylpyrrolidone)-Modified Silicas

“…This protein-coated polymer provides a novel matrix for covalent immobilization of enzymes. 20 Also crosslinkage allows the synthesis of various materials for immunosorbent assays. 21 The last two methods of protein immobilization are not always suitable for immobilization of all proteins because of the hard synthetic conditions, which lead to protein denaturation.…”

Urea Biosensor Based on a CO2 Microsensor

“…Covalent attachment and cross-linking can prevent leaching and be effective and durable, but there is a high risk of decreasing the enzyme activity by disturbed enzyme conformation . The addition of compounds with suitable functional groups for covalent bonding has been proposed to lower the detrimental effects on enzyme activity by lowering the covalent bonding directly to the enzyme. , Support protein such as BSA or polymers such as chitosan and alginate are used for cross-linking to enzymes by the use of glutaraldehyde, , and for urease, it was shown to result in a retained activity of more than 50% as compared to the free enzyme, with some activity still being present after a few weeks …”

Protein-Based 3D Microstructures with Controllable Morphology and pH-Responsive Properties

“…Native proteins have congenital advantages, such as nontoxicity, biodegradability, and biocompatibility, − which prompt the diverse applications considering the chemical, optical, mechanical, and electromagnetic properties. − Bovine serum albumin (BSA) as one of the native proteins has been widely studied due to its low cost, stimuli-responsive property, and similar nature to human serum albumin. − For example, it can be used as a carrier of ions, metabolites, drugs, and hormones, ,, as well as stabilizing and immobilizing agent for some enzymes. , Most notably, BSA-based tunable, dynamic, and responsive “smart” three-dimensional (3D) microstructures with well-defined geometries and repeatability are crucial for regenerative medicine and tissue engineering. Two-photon polymerization (TPP) microfabrication technology has excellent capability to produce 3D microstructures with high precision, ,− which is promising for achieving the BSA microstructure with precise configuration.…”

“…8−13 For example, it can be used as a carrier of ions, metabolites, drugs, and hormones, 7,14,15 as well as stabilizing and immobilizing agent for some enzymes. 16,17 Most notably, BSA-based tunable, dynamic, and responsive "smart" three-dimensional (3D) microstructures with well-defined geometries and repeatability are crucial for regenerative medicine and tissue engineering. Two-photon polymerization (TPP) microfabrication technology has excellent capability to produce 3D microstructures with high precision, 4,18−20 which is promising for achieving the BSA microstructure with precise configuration.…”