Construction of a selective cleavage system for a protein displayed on the cell surface of yeast |
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Authors: | Michiko Kato Haruko Maeda Masayuki Kawakami Seizaburo Shiraga Mitsuyoshi Ueda |
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Institution: | (1) Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku Kyoto, 606-8502, Japan;(2) Advanced Core Technology Laboratories, Fuji Photo Film Co., Ltd., 210 Nakamura, Minami-Ashigara, Kanagawa 250-0193, Japan |
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Abstract: | We constructed a novel protein-purification system in which Saccharomyces cerevisiae with a protein displayed on the cell surface is harvested and the displayed protein is then cleaved from the cell surface. GFPuv was used as a model protein in this cell surface engineering experiment. In this system, the C-terminal 320 amino acids of α-agglutinin were bound to the C-terminal of GFPuv for display on the cell surface. In this novel system, the insertion of the recognition sequence-encoding gene of protease factor Xa between GFPuv and α-agglutinin was successfully carried out. The GFPuv, displayed by the insertion, was successfully cleaved from yeast cell surface by treatment with factor Xa, and could be easily recovered. By removing such a protease with well-known properties, the displayed protein could be isolated and purified with relative ease. |
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