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.
Keywords: 10th human fibronectin type III domain; Autotransporter; Cell surface display; Cold-active esterase; Permafrost; Psychrobacter cryohalolentis K5T.