Efficient expression of full-length antibodies in the cytoplasm of engineered bacteria
Open accessAbstract
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.
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E. coli SHuffle®T7 Express is useful for the expression of complex proteins containing disulfide bonds.
“The usefulness of E. coli SHuffle®T7 Express for the expression of complex proteins containing disulfide bonds has already been demonstrated [6, 22, 23].”
From High level in vivo mucin-type glycosylation in Escherichia coli (2018) · cites this paper as [22]
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- Authors
- Robinson, M.-P., Ke, N., Lobstein, J., Peterson, C., Szkodny, A., Mansell, T. J., Tuckey, C., Riggs, P. D., Colussi, P. A., Noren, C. J., Taron, C. H., DeLisa, M. P., & Berkmen, M.
- Affiliations
- Cornell University (US)
- New England Biolabs (United States) (US)
- Corresponding author
- DeLisa, Matthew P.
- Berkmen, Mehmet
- Venue
- Nature Communications
- Published
- 2015-08-27
- Volume
- 6
- Issue
- 1
- Pages
- 8072-8072
- ISSN
- 2041-1723 (ISSN-L)
- PMC
- PMC4560801
- OpenAlex
- W1239799147
- Cited by
- 131
- References
- 52
- Citations by year
- 2026: 5 · 2025: 13 · 2024: 6 · 2023: 13 · 2022: 11 · 2021: 17 · 2020: 20 · 2019: 10 · 2018: 12 · 2017: 15 · 2016: 7 · 2015: 1
- FWCI
- 9.44
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Locations
- Nature Communications (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- RePEc: Research Papers in Economics (submittedVersion)
- Funders
- National Science Foundation
- National Institutes of Health (1r41ai092969-01a1)
- National Institute of Allergy and Infectious Diseases (AI092969-01A1, R41 AI092969)
- Topics
- Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Monoclonal and Polyclonal Antibodies Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein purification and stability
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Microbial Metabolic Engineering and Bioproduction
- Keywords
- IgG, Escherichia coli, disulfide bond stability, Fcγ receptor binding, effector function, membrane translocation
- MeSH terms
- Antibodies, Antibody Formation, Bacteriophages, Cytoplasm, Electrophoresis, Polyacrylamide Gel, Enzyme-Linked Immunosorbent Assay, Escherichia coli, Glycosylation, Immunoglobulin G, Plasmids, Blotting, Western, Surface Plasmon Resonance, Protein Transport, Organisms, Genetically Modified
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