Premium
Escherichia coli inner membrane display system for high‐throughput screening of dimeric proteins
Author(s) -
Jo Migyeong,
Hwang Bora,
Yoon Hyun Woung,
Jung Sang Taek
Publication year - 2018
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.26826
Subject(s) - escherichia coli , throughput , inner membrane , chemistry , membrane protein , high throughput screening , escherichia coli proteins , biophysics , biochemistry , membrane , microbiology and biotechnology , computational biology , biology , gene , computer science , telecommunications , wireless
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.