Large-scale separation and production of engineered proteins,designed for facilitated recovery in detergent-based aqueous two-phase extraction systems |
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| Affiliation: | 1. Institute of Enzyme Technology, Heinrich-Heine University, Düsseldorf, D-52426 Juelich, Germany;2. Department of Biochemistry, Center for Chemistry and Chemical Engineering, Lund University, P.O. Box 124, S-22100 Lund, Sweden;3. VTT Biotechnology and Food Research, P.O. Box 1500, FIN-02044 VTT, Espoo, Finland;4. Genencor International, B.V., Archimedesweg 30, 2333 CN Leiden, The Netherlands;1. Laboratory of Inorganic Chemistry, Faculty of Sciences, BP 1171, 3000 Sfax, University of Sfax, Tunisia;2. Laboratory of Solid States, Department of Physics, Faculty of Sciences, BP 1171, 3000 Sfax, University of Sfax, Tunisia;1. Department of Molecular Medicine and Bioprocesses, Biotechnology Institute, National University of Mexico (UNAM), Mexico;2. Laboratory of Cellular Neuropharmacology, Biophysics and Biochemistry Center, Instituto Venezolano de Investigaciones Científicas (IVIC), Caracas, Venezuela;3. Histology Service, Biophysics and Biochemistry Center, Instituto Venezolano de Investigaciones Cientificas (IVIC), Caracas, Venezuela;1. Department of Chemical Engineering, Universidade de Vigo, P.O. Box 36310, Vigo, Spain;2. Design in Engineering Department, Universidade de Vigo, P.O. Box 36310, Vigo, Spain |
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| Abstract: | The feasibility and scalability of extraction in detergent-based aqueous two-phase systems for the separation of proteins from culture broth is demonstrated. At the same time the large-scale production of a fusion protein and the influence of cultivation scale on the efficiency of separation were investigated. An amphiphilic fusion protein EGIcore-HFBI was chosen, consisting of the catalytic core of the cellulase endoglucanase I and the small protein hydrophobin I, expressed homologously in Trichoderma reesei. Using the technical nonionic detergent Agrimul NRE 1205 the separation was successfully scaled up to 1200 l. No differences in yield or in partition coefficient were observed at 10 ml and 1200 l scale. Changes in the fermentation temperature and scale, however, can influence the properties of the protein and thus alter partition coefficient and yield. The decreased separation efficiency appears to be correlated with changes in glycosylation at lower cultivation temperatures. |
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