Direct affinity immobilization of recombinant heparinase I fused to maltose binding protein on maltose-coated magnetic nanoparticles |
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| Institution: | 1. Key Laboratory for Industrial Biocatalysis, Ministry of Education, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Haidian, Beijing 100084, PR China;2. Key Lab of Organic Optoelectronic & Molecular Engineering, Department of Chemistry, Tsinghua University, Haidian, Beijing 100084, PR China;1. School of Health and Biomedical Sciences, RMIT University, Victoria, 3083, Australia;2. School of Chemistry, The University of Melbourne, Victoria, 3010, Australia;3. Alfred Hospital Radiation Oncology, Melbourne, Australia;4. Dept. of Clinical Radiology, Hiroshima Int. University, Hiroshima, 739-452, Japan;1. Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, PR China;2. State Key Laboratory of Chemical Engineering, Tianjin University, Tianjin 300072, PR China;3. Tianjin Key Laboratory of Membrane Science and Desalination Technology, Tianjin University, Tianjin 300072, PR China;4. The Co-Innovation Center of Chemistry and Chemical Engineering of Tianjin, Tianjin 300072, PR China;1. Joint Graduate School for Energy and Environment (JGSEE), King Mongkut''s University of Technology Thonburi, Bangmod, Bangkok 10140, Thailand;2. Enzyme Technology Laboratory, National Center for Genetic Engineering and Biotechnology, 113 Thailand Science Park, Phahonyothin Road, Khlong Nueng, Khlong Luang, Pathum Thani 12120, Thailand;1. Atmosphere and Ocean Research Institute, The University of Tokyo, Chiba, Japan;2. Department of Earth and Planetary Science, The University of Tokyo, Tokyo, Japan;3. Research Institute for Global Change, Japan Agency for Marine-Earth Science and Technology, Yokosuka, Japan;4. Institute of Low Temperature Science, Hokkaido University, Sapporo, Japan;5. Far Eastern Regional Hydrometeorological Research Institute, Vladivostok, Russia |
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| Abstract: | Hybrid magnetic Fe3O4@SiO2-poly(ethylene oxide)-maltose (Fe3O4@SiO2-PEO-mal) nanoparticles synthesized by our group can be used as affinity adsorption carriers for direct separation of maltose binding protein-fused Hep I (MBP-Hep I) from a crude enzyme solution in a magnetic field. In this work, different PEO molecular weights for Fe3O4@SiO2-PEO-mal nanoparticles were used for characterizing of MBP-Hep I immobilization. The results showed that all four kinds of Fe3O4@SiO2-PEO-mal magnetic nanoparticles (6k, 20k, 35k and 100k for PEO) exhibited excellent adsorption capacities and the adsorption ratio increased as the PEO molecular weight increased from 6k to 100k. All four kinds of immobilized MBP-Hep I exhibited significantly improved stability at 30 °C compared with free MBP-Hep I and their half-lives were 20–50 times that of the free MBP-Hep I. Fe3O4@SiO2-PEO-mal nanoparticles with a PEO molecular weight of 100k were best able to immobilize MBP-Hep I (Fe3O4@SiO2-PEO100k-mal-MBP-Hep I). The molecular weight distribution profiles and anticoagulant activities, obtained from heparin depolymerization by free Hep I, free MBP-Hep I and Fe3O4@SiO2-PEO100k-mal-MBP-Hep I were the same. Furthermore, Fe3O4@SiO2-PEO100k-mal-MBP-Hep I exhibited reasonable reusability during enzymatic production of low molecular weight heparins (LMWHs). |
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| Keywords: | Affinity Adsorption Heparinase I Immobilization Magnetic nanoparticles Enzyme biocatalysis |
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