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Cell-free production of functional antibody fragments

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Abstract

Botulinum neurotoxin serotype B (BoNT/B)-specific Fab was expressed in a cell-free protein synthesis system derived from an E. coli extract. The cell-free synthesized antibody fragment was found to be effective in neutralizing the toxicity of BoNT/B in animal studies. Expression of functional Fab required an appropriately controlled and stably maintained redox potential. Under an optimized redox condition, the cell extract, whose disulfide reducing activity had been exhausted, could generate bio-functional Fab molecules. Use of a cell extract enriched with molecular chaperones (GroEL/ES) and disulfide bond isomerases were effective in obtaining larger quantities of functional Fab. Under the optimized reaction conditions, approximately 30 μg of functional Fab was obtained after purification from 1 mL reaction mixture.

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Acknowledgment

This work was supported by a grant from the Basic Research Program of the Korea Science & Engineering Foundation (Grant No. R01-2007-000-20980-0).

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Authors and Affiliations

  1. Institute of Molecular Biology and Genetics, Seoul National University, Seoul, Korea

    In-Seok Oh

  2. Abxign, Inc., Samsung Cancer Research Building, Seoul, Korea

    Ji-Chul Lee

  3. Department of Microbiology and Immunology, Eulji University School of Medicine, Taejon, Korea

    Myung-shin Lee

  4. Department of Biochemistry and Molecular Biology, Cancer Research Institute, Seoul National University College of Medicine, Seoul, South Korea

    Jun-ho Chung

  5. Department of Fine Chemical Engineering and Applied Chemistry, Chungnam National University, Taejon, 305-764, Korea

    Dong-Myung Kim

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  1. In-Seok Oh
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  2. Ji-Chul Lee
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  3. Myung-shin Lee
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  4. Jun-ho Chung
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  5. Dong-Myung Kim
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Correspondence to Dong-Myung Kim.

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Oh, IS., Lee, JC., Lee, Ms. et al. Cell-free production of functional antibody fragments. Bioprocess Biosyst Eng 33, 127–132 (2010). https://doi.org/10.1007/s00449-009-0372-3

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