Stability enhancement of an O2-tolerant NAD+-reducing [NiFe]-hydrogenase by a combination of immobilisation and chemical modification |
| |
| Affiliation: | 1. TU Berlin, Institut für Chemie, Fachgebiet Technische Chemie/Enzymtechnologie, Str. des 17. Juni 124, 10623 Berlin, Germany;2. University of Oxford, Department of Chemistry, Inorganic Chemistry Laboratory, South Parks Road, Oxford OX1 3QR, UK;3. Humboldt-Universität zu Berlin, Institut für Biologie/Mikrobiologie, Chausseestraße 117, 10115 Berlin, Germany;1. Department of Chemistry, University of Torino, via P. Giuria 7, 10125 Torino, Italy;2. NIS – Nanostructured Interfaces and Surfaces and “G. Scansetti” Interdepartmental Centre for Studies on Asbestos and Other Toxic Particulate, University of Torino, Italy;3. Department of Drug Science and Technology, University of Torino, via P. Giuria 7, 10125 Torino, Italy;1. Department of Cell and Molecular Physiology, Loyola University Chicago, 2160 South First Avenue, Maywood, IL 60153, United States;2. Department of Molecular Biophysics and Physiology, Rush University Medical Center, 1750 West Harrison Street, Chicago, IL 60612, United States;1. Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan;2. Division of Applied Life Sciences, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Naka-ku, Sakai 559-8531, Japan;3. Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto 606-8502, Japan;4. Research & Development Center, Nagase & Co., Ltd., 2-2-3 Muratani, Nishi-ku, Kobe 651-2241, Japan;5. Faculty of Bioenvironmental Science, Kyoto Gakuen University, Sogabe-cho, Kameoka 621-8555, Japan;1. Department of Food, Environmental and Nutritional Sciences – DeFENS, University of Milan, via Mangiagalli 25, 20133 Milano, Italy;2. Department of Pharmaceutical Sciences – DISFARM, University of Milan, via Mangiagalli 25, 20133 Milano, Italy;1. Department of Physics, Emory University, Atlanta, Georgia;1. Chair of Chemistry of Biogenic Resources, Technical University of Munich; Campus Straubing, 94315 Straubing, Germany;2. SynBioFoundry@TUM, Technical University of Munich, 94315 Straubing, Germany;3. School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, QLD 4072, Australia |
| |
| Abstract: | The oxygen-tolerant, NAD+-reducing soluble hydrogenase (SH) from Ralstonia eutropha H16 is a promising catalyst for cofactor regeneration in enzyme-catalysed reduction processes. The technical use of the isolated enzyme, however, is limited by its relatively low stability under operational conditions such as agitation, elevated temperature or addition of co-solvents. The maximum half-life at a reaction temperature of 35 °C and pH 8.0 was only 5.3 h. In order to enhance the stability of the enzyme, it was immobilised onto the anionic resin Amberlite™ FPA54. At an immobilisation yield of 93.4% for adsorptive and 100% for covalent attachment, corresponding activities of 48.9 and 39.3%, respectively, were obtained. Covalent binding always yielded superior stabilisation. At elevated temperature and under agitation, stabilisation was further increased by modification of the covalently bound SH with methoxy-poly(ethylene) glycol (mPEG). A comparable effect was not achieved when SH modification was performed before immobilisation. In stationary aqueous solution, half-lives of up to 161 h at 25 °C and 32 h at 35 °C were obtained. In presence of the technically relevant co-solvents DMSO, DMF, 2-propanol and [EMIM][EtSO4] half-lives of 14–29 h can now be achieved. |
| |
| Keywords: | Bidirectional [NiFe]-hydrogenase Immobilisation Amberlite™ Chemical modification |
| 本文献已被 ScienceDirect 等数据库收录！ |
|
