title: Covalent Immobilization of Catalase onto Regenerated Silk Fibroins via Tyrosinase-Catalyzed Cross-Linking.
abstract: Regenerated silk fibroins could be used as medical scaffolds and carrier materials for enzyme immobilization. In the present work, tyrosinase enzyme was used for enzymatic oxidation of silk fibroins, followed by immobilization of catalase onto the fibroin surfaces through physical adsorption and covalent cross-linking as well. Spectrophotometry, SDS-PAGE, and Fourier transform infrared spectroscopy (FTIR) were used to examine the efficiency of enzymatic oxidation and catalase immobilization, respectively. The results indicate that tyrosine residues in silk fibroins could be oxidized and converted to the active o-quinones. Incubating silk fibroins with catalase and tyrosinase led to a noticeable change of molecular weight distribution, indicating the occurrence of the cross-links between silk fibroins and catalase molecules. Two different pathways were proposed for the catalase immobilizations, and the method based on grafting of catalase onto the freeze-dried fibroin membrane is more acceptable. The residual enzyme activity for the immobilized catalase exhibited higher than that of the control after repeated washing cycles. Meanwhile, the thermal stability and alkali resistance were also slightly improved as compared to free catalase. The mechanisms of enzymatic immobilization are also concerned.
authors.affiliation: Key Laboratory of Science and Technology of Eco-Textile, Ministry of Education, Jiangnan University, Wuxi, 214122, People's Republic of China, pwang@jiangnan.edu.cn.
authors.forename: Ping
authors.initials: P
authors.lastname: Wang
authors.forename: Chenglong
authors.initials: C
authors.lastname: Qi
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Yu
authors.forename: Jiugang
authors.initials: J
authors.lastname: Yuan
authors.forename: Li
authors.initials: L
authors.lastname: Cui
authors.forename: Gengtie
authors.initials: G
authors.lastname: Tang
authors.forename: Qiang
authors.initials: Q
authors.lastname: Wang
authors.forename: Xuerong
authors.initials: X
authors.lastname: Fan
mesh_terms: D000818:Animals; D055162:Biocatalysis; D002374:Catalase / Q000378:metabolism*; D002417:Cattle; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents / Q000378:metabolism*; D004295:Dihydroxyphenylalanine / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000378:metabolism*; D005349:Fibroins / Q000737:chemistry* / Q000378:metabolism; D005612:Freeze Drying; D006863:Hydrogen-Ion Concentration; D014442:Monophenol Monooxygenase / Q000378:metabolism*; D010084:Oxidation-Reduction; D012996:Solutions; D013056:Spectrophotometry, Ultraviolet; D013696:Temperature
chemical_list: D003432:Cross-Linking Reagents; D004800:Enzymes, Immobilized; D012996:Solutions; D004295:Dihydroxyphenylalanine; D005349:Fibroins; D002374:Catalase; D014442:Monophenol Monooxygenase