Isolation of full-length IgG antibodies from combinatorial libraries expressed in the cytoplasm of Escherichia coli

Michael-Paul Robinson, Jinjoo Jung, Natalia Lopez-Barbosa, Matthew Chang, Mingji Li, Thapakorn Jaroentomeechai, Emily C. Cox, Xiaolu Zheng, Mehmet Berkmen and Matthew P. DeLisa ()
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Michael-Paul Robinson: Cornell University
Jinjoo Jung: Cornell University
Natalia Lopez-Barbosa: Cornell University
Matthew Chang: Cornell University
Mingji Li: Cornell University
Thapakorn Jaroentomeechai: Cornell University
Emily C. Cox: Cornell University
Xiaolu Zheng: Cornell University
Mehmet Berkmen: New England Biolabs
Matthew P. DeLisa: Cornell University

Nature Communications, 2023, vol. 14, issue 1, 1-12

Abstract: Abstract Here we describe a facile and robust genetic selection for isolating full-length IgG antibodies from combinatorial libraries expressed in the cytoplasm of redox-engineered Escherichia coli cells. The method is based on the transport of a bifunctional substrate comprised of an antigen fused to chloramphenicol acetyltransferase, which allows positive selection of bacterial cells co-expressing cytoplasmic IgGs called cyclonals that specifically capture the chimeric antigen and sequester the antibiotic resistance marker in the cytoplasm. The utility of this approach is first demonstrated by isolating affinity-matured cyclonal variants that specifically bind their cognate antigen, the leucine zipper domain of a yeast transcriptional activator, with subnanomolar affinities, which represent a ~20-fold improvement over the parental IgG. We then use the genetic assay to discover antigen-specific cyclonals from a naïve human antibody repertoire, leading to the identification of lead IgG candidates with affinity and specificity for an influenza hemagglutinin-derived peptide antigen.

Date: 2023
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DOI: 10.1038/s41467-023-39178-x

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