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

Author

Listed:
  • 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
    Cornell University
    Cornell University)

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.

Suggested Citation

  • Michael-Paul Robinson & Jinjoo Jung & Natalia Lopez-Barbosa & Matthew Chang & Mingji Li & Thapakorn Jaroentomeechai & Emily C. Cox & Xiaolu Zheng & Mehmet Berkmen & Matthew P. DeLisa, 2023. "Isolation of full-length IgG antibodies from combinatorial libraries expressed in the cytoplasm of Escherichia coli," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-39178-x
    DOI: 10.1038/s41467-023-39178-x
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    References listed on IDEAS

    as
    1. Michael-Paul Robinson & Na Ke & Julie Lobstein & Cristen Peterson & Alana Szkodny & Thomas J. Mansell & Corinna Tuckey & Paul D. Riggs & Paul A. Colussi & Christopher J. Noren & Christopher H. Taron &, 2015. "Efficient expression of full-length antibodies in the cytoplasm of engineered bacteria," Nature Communications, Nature, vol. 6(1), pages 1-9, November.
    2. Bunyarit Meksiriporn & Morgan B. Ludwicki & Erin A. Stephens & Allen Jiang & Hyeon-Cheol Lee & Dujduan Waraho-Zhmayev & Lutz Kummer & Fabian Brandl & Andreas Plückthun & Matthew P. DeLisa, 2019. "A survival selection strategy for engineering synthetic binding proteins that specifically recognize post-translationally phosphorylated proteins," Nature Communications, Nature, vol. 10(1), pages 1-10, December.
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