Simple Purification of a Foreign Protein Using Polyhedrin Fusion in a Baculovirus Expression System

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概要

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• Previously, we found that expression by translational fusion of the polyhedrin (Polh)-green fluorescence protein (GFP) led to the formation of granular structures, and that these fluorescent granules were easily precipitated by high-speed centrifugation. Here, we developed an easy, fast, mass purification system using this baculovirus expression system (BES). An enhanced GFP (EGFP) fused with the Polh gene at the N-terminus, including a linker and enterokinase (EK) site between Polh and EGFP, was expressed in Sf9 cells. The cells infected by AcPolhEKA-EGFP produced fluorescent granules. The EGFP fusion protein was purified from granule-containing cells in three steps: cell harvest, sonication, and EK digestion. Through final enterokinase digestion, EGFP presented mainly in the supernatant, and this supernatant fraction also showed a pure EGFP band. These results suggest that a combined procedure of Polh fusion expression and enterokinase digestion can be used for rapid and easy purification of other proteins.

著者

• Roh Jong

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

• Choi Jae

  Research Institute for Agriculture and Life Sciences, Seoul National University

• Kang Joong

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

• WANG Yong

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

• SHIM Hee

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

• LIU Qin

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

• TAO Xueying

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

• XU Hong

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

• HYUN Jin-Ho

  Department of Biosystems and Biomaterials Science and Engineering, College of Agriculture and Life S

• WOO Soo

  Department of Plant Medicine, College of Agriculture, Life and Environment Sciences, Chungbuk Nation

• JIN Byung

  College of Natural Resources and Life Science, Dong-A University

• JE Yeon

  Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National U

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