Efficient expression of full-length antibodies in the cytoplasm of engineered bacteria

• Published in: Nature communications Vol. 6; no. 1; p. 8072
• Main Authors: Robinson, Michael-Paul, Ke, Na, Lobstein, Julie, Peterson, Cristen, Szkodny, Alana, Mansell, Thomas J., Tuckey, Corinna, Riggs, Paul D., Colussi, Paul A., Noren, Christopher J., Taron, Christopher H., DeLisa, Matthew P., Berkmen, Mehmet
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 27.08.2015; Nature Publishing Group; Nature Pub. Group
• Subjects: 631/1647/1511; 631/326/41; 82/1; 82/103; 82/16; 82/29; 82/80; 82/83; Antibodies; Antibody Formation - genetics; Bacteriophages - genetics; Blotting, Western; Cytoplasm - metabolism; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Escherichia coli - genetics; Glycosylation; Humanities and Social Sciences; Immunoglobulin G - biosynthesis; multidisciplinary; Organisms, Genetically Modified - genetics; Plasmids - genetics; Protein Transport; Science; Science (multidisciplinary); Surface Plasmon Resonance
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/ncomms9072

Current methods for producing immunoglobulin G (IgG) antibodies in engineered cells often require refolding steps or secretion across one or more biological membranes. Here, we describe a robust expression platform for biosynthesis of full-length IgG antibodies in the Escherichia coli cytoplasm. Synthetic heavy and light chains, both lacking canonical export signals, are expressed in specially engineered E. coli strains that permit formation of stable disulfide bonds within the cytoplasm. IgGs with clinically relevant antigen- and effector-binding activities are readily produced in the E. coli cytoplasm by grafting antigen-specific variable heavy and light domains into a cytoplasmically stable framework and remodelling the fragment crystallizable domain with amino-acid substitutions that promote binding to Fcγ receptors. The resulting cytoplasmic IgGs—named ‘cyclonals’—effectively bypass the potentially rate-limiting steps of membrane translocation and glycosylation. Current methods for production of monoclonal antibodies often require refolding steps or secretion across biological membranes. Here, Robinson et al. describe engineered E. coli strains for efficient production of functional immunoglobulin G antibodies in the bacterial cytoplasm.