9MVX

Crystal structure of knob-in-hole immunoglobulin G1 Fc heterodimer with P374A

Summary for 9MVX

• Entry DOI: 10.2210/pdb9mvx/pdb
• Descriptor: Isoform 1 of Immunoglobulin heavy constant gamma 1 HC1 (Hole), Isoform 1 of Immunoglobulin heavy constant gamma 1 HC2 (Knob), 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[2-acetamido-2-deoxy-beta-D-glucopyranose-(1-2)-alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose, ... (5 entities in total)
• Functional Keywords: human fc fragment, immune response, immune system, knob, hole, knob-in-hole, kih, proala, proline-to-alanine, bispecific antibody, bsab, bsigg, cis-peptide
• Biological source: Homo sapiens (human); More
• Total number of polymer chains: 2
• Total formula weight: 52954.36

Authors

Choi, W.S.,Tilegenova, C.,Are, M.,Zwolak, A.,Shaffer, P.,Sharma, S. (deposition date: 2025-01-16, release date: 2025-11-12)

Primary citation

Tilegenova, C.,Liu, T.,Zhao, Q.,Are, M.,Zhao, Y.,Choi, W.S.,Bhaumik, A.,Steele, R.,Manieri, N.A.,Turegun, B.,Ni, A.,Cardoso, R.M.F.,Shaffer, P.,Clark, D.,Ernst, R.,Li, W.,Taylor, T.,Swaminathan, S.K.,Ramaraju, B.,Liaw, K.,Jacobs, S.A.,Sharma, S.,Cheung, W.C.,Zwolak, A.
Folding-mediated secretion of pure bispecific antibodies.
Nat.Biotechnol., 2025

PubMed Abstract: Bispecific antibodies (bsAbs) can enable therapeutic mechanisms, such as dual antigen targeting or receptor agonism, that are impossible using monoclonal antibodies. BsAbs with IgG-like format (bsIgG) are comprised of two unique heavy chains, each having a cognate light chain. Co-expression of these four unique polypeptides often leads to several mispaired species that are difficult to separate from the target bsIgG due to their similar biophysical properties. Here we describe a set of mutations called ProAla that exploit a the unfolded protein response pathway of cells. ProAla heavy chains are engineered with higher folding energy barriers such that only the cognate light and heavy chains can induce folding, chaperone release and secretion. The structures of the ProAla Fab and Fc regions are identical in structure to normal antibodies, enabling maintenance of half-life and function. Mispaired polypeptides fail to secrete from the cell due to enhanced interaction with the endoplasmic reticulum chaperone BiP, resulting in increased purity of secreted bsIgGs.

PubMed: 41057658
DOI: 10.1038/s41587-025-02842-2

Experimental method

X-RAY DIFFRACTION (1.84 Å)