title: Cell-free production and streamlined assay of cytosol-penetrating antibodies.
abstract: Antibodies that target intracellular proteins hold great promise in the development of novel therapeutic interventions for various diseases. In particular, antibodies that can cross cellular membranes have potential applications in controlling disease-related intracellular protein-protein interactions. Given the large number of cytosolic proteins and complicated interactions that are potentially involved in disease development, discovery of antibodies targeting intracellular proteins requires iterative cycles of expression and assessment of candidate antibodies. Because current cell-based expression methods do not provide sufficient throughput for production and assay of cytosol-penetrating antibodies, we integrated a cell-free protein synthesis system designed to provide optimal conditions for production of functional antibodies with a cytosol-penetration assay. The proposed approach of consolidating cell-free synthesis and cell-based assay will substantially expand the capability of discovering and engineering antibodies that can cross the cell membrane and effectively control protein-mediated cellular functions. Biotechnol. Bioeng. 2016;113: 2107-2112. © 2016 Wiley Periodicals, Inc.
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, Chungnam National University, Daejeon, 34134, Korea.
authors.forename: Seung Eui
authors.initials: SE
authors.lastname: Min
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, Chungnam National University, Daejeon, 34134, Korea.
authors.forename: Kyung-Ho
authors.initials: KH
authors.lastname: Lee
authors.affiliation: Department of Molecular Science and Technology, Ajou University, Suwon, Korea.
authors.forename: Seong-Wook
authors.initials: SW
authors.lastname: Park
authors.affiliation: Department of Molecular Science and Technology, Ajou University, Suwon, Korea.
authors.forename: Tae Hyeon
authors.initials: TH
authors.lastname: Yoo
authors.affiliation: Department of Bio and Brain Engineering, Korea Advanced Institute of Science and Technology, Daejeon, Korea.
authors.forename: Chan Hee
authors.initials: CH
authors.lastname: Oh
authors.affiliation: Department of Bio and Brain Engineering, Korea Advanced Institute of Science and Technology, Daejeon, Korea.
authors.forename: Ji-Ho
authors.initials: JH
authors.lastname: Park
authors.affiliation: Department of Polymer Science and Engineering, Chungnam National University, Daejeon, Korea.
authors.forename: Sung Yun
authors.initials: SY
authors.lastname: Yang
authors.affiliation: Department of Molecular Science and Technology, Ajou University, Suwon, Korea.
authors.forename: Yong-Sung
authors.initials: YS
authors.lastname: Kim
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, Chungnam National University, Daejeon, 34134, Korea. dmkim@cnu.ac.kr.
authors.forename: Dong-Myung
authors.initials: DM
authors.lastname: Kim
mesh_terms: D000911:Antibodies, Monoclonal / Q000235:genetics / Q000378:metabolism*; D002474:Cell-Free System / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D006367:HeLa Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D015202:Protein Engineering / Q000379:methods*
chemical_list: D000911:Antibodies, Monoclonal
keywords: antibody; cell-free protein synthesis; cytosol-penetrating antibody; protein-protein interaction; therapeutic