Multimer formation is indispensable to the intrinsicbiologicalfunctions of many natural proteins. For example, the human immunoglobulin G (IgG) antibody has two variable regions (heavy chain variable domain [VH] and light chain variable domain [VL]) that must be assembled for specific antigen binding, and homodimerization of the antibody's Fc domain is essential for eliciting therapeutic effector functions. For the more efficient high-throughput directed evolution of multimeric proteins with ease of cultivation and handling, here we report a membrane protein drift and assembly (MPDA) system, in which a multimeric protein is displayed on a bacterial inner membrane by drifting and auto-assembling membrane-anchored subunit polypeptides. This system enabled the auto-assembly of membrane-tethered Fv domains (VH and VL) or the monomeric Fc domain into a functional hetero- or homodimeric protein complex on the bacterial inner membrane. This system could also be used to enrich a desired engineered Fc variant from a mixture containing a million-fold excess of wild-type Fc domain, indicating the applicability of the MPDA system for the high-throughput directed evolution of a variety of multimeric proteins, such as cytokines, enzymes, or structural proteins.
Bibliographical noteFunding Information:
The Bio & Medical Technology Development Program through the National Research Foundation of Korea funded by the Ministry of Science, ICT, and Future Planning, Grant/ Award Number: 2017M3A9C8060552; The Basic Science Research Program through the National Research Foundation of Korea funded by the Ministry of Science, ICT, and Future Planning, Grant/Award Number: 2016R1C1B2007434; The Pioneer Research Center Program through the National Research Foundation of Korea funded by the Ministry of Science, ICT, and Future Planning, Grant/Award Number: 2014M3C1A3051460
© 2018 Wiley Periodicals, Inc.
- bacterial display
- directed evolution
- library screening
- multimeric protein
ASJC Scopus subject areas
- Applied Microbiology and Biotechnology