Fusion with the cold-active esterase facilitates autotransporter-based surface display of the 10th human fibronectin domain in Escherichia coli
| Autor: | S. Sh. Gapizov, K. A. Novototskaya-Vlasova, Elizaveta Rivkina, A. V. Zlobinov, Dmitry A. Dolgikh, Mikhail P. Kirpichnikov, Shingarova Ln, E. F. Boldyreva, Lada E. Petrovskaya |
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| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Scaffold protein Type V Secretion Systems 030106 microbiology Fibronectin type III domain medicine.disease_cause Microbiology Esterase 03 medical and health sciences medicine Escherichia coli Humans biology Chemistry Cell Membrane Esterases Psychrobacter General Medicine Psychrobacter cryohalolentis biology.organism_classification Fusion protein Recombinant Proteins Cell biology Fibronectins Fibronectin Cold Temperature 030104 developmental biology biology.protein Molecular Medicine Linker |
| Zdroj: | Extremophiles : life under extreme conditions. 22(1) |
| ISSN: | 1433-4909 |
| Popis: | Cell surface display is a popular approach for the construction of whole-cell biocatalysts, live vaccines, and screening of combinatorial libraries. To develop a novel surface display system for the popular scaffold protein 10th human fibronectin type III domain (10Fn3) in Escherichia coli cells, we have used an α-helical linker and a C-terminal translocator domain from previously characterized autotransporter from Psychrobacter cryohalolentis K5T. The level of 10Fn3 passenger exposure at the cell surface provided by the hybrid autotransporter Fn877 and its C-terminal variants was low. To improve it, the fusion proteins containing 10Fn3 and the native autotransporter passenger Est877 or the cold-active esterase EstPc in different orientations were constructed and expressed as passenger domains. Using the whole-cell ELISA and activity assays, we have demonstrated that N-terminal position of EstPc in the passenger significantly improves the efficiency of the surface display of 10Fn3 in E. coli cells. |
| Databáze: | OpenAIRE |
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