An antibody with Fab-constant domains exchanged for a pair of CH3 domains
| Autor: | Florian Rüker, Mathias Gotsmy, Gordana Wozniak-Knopp, Jan Walther Perthold, Katharina Stadlbauer, Gerhard Stadlmayr, Stefan Becker |
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| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Models Molecular Receptor ErbB-2 Thermal Stability lcsh:Medicine NK cells Protein Engineering Biochemistry Database and Informatics Methods Trastuzumab Cellular types Amino Acids lcsh:Science Antibody-dependent cell-mediated cytotoxicity Staining Multidisciplinary biology Chemistry Organic Compounds Physics Immune cells Cell Staining Antibodies Monoclonal Recombinant Proteins Killer Cells Natural Physical Sciences Thermodynamics White blood cells Antibody Immunoglobulin Constant Regions Sequence Analysis medicine.drug Research Article Cell Binding Cell Physiology Blood cells Bioinformatics Materials Science Material Properties Immunology Research and Analysis Methods Cell Line 03 medical and health sciences Immunoglobulin Fab Fragments Protein Domains Sequence Motif Analysis medicine Genetics Point Mutation Sulfur Containing Amino Acids Animals Humans Cysteine Medicine and health sciences Reporter gene 030102 biochemistry & molecular biology Point mutation Mutagenesis Organic Chemistry Receptors IgG lcsh:R Chemical Compounds Antibody-Dependent Cell Cytotoxicity Biology and Life Sciences Proteins Cell Biology 030104 developmental biology Solubility Animal cells Cell culture Specimen Preparation and Treatment Immunoglobulin G Mutation Biophysics biology.protein Mutagenesis Site-Directed lcsh:Q |
| Zdroj: | PLoS ONE, Vol 13, Iss 4, p e0195442 (2018) PLoS ONE |
| ISSN: | 1932-6203 |
| Popis: | We have designed a complete antibody-like construct where the CH1 and Cκ domains are exchanged for a pair of the CH3 domains and efficient pairing of the heavy and light variable domain is achieved using “Knobs-into-Holes” strategy. This construct, composed of only naturally occurring immunoglobulin sequences without artificial linkers, expressed at a high level in mammalian cells, however exhibited low solubility. Rational mutagenesis aimed at the amino acid residues located at the interface of the variable domains and the exchanged CH3 domains was applied to improve the biophysical properties of the molecule. The domain-exchanged construct, including variable domains of the HER2/neu specific antibody trastuzumab, was able to bind to the surface of the strongly HER2/neu positive cell line SK-BR3 4-fold weaker than trastuzumab, but could nevertheless incite a more potent response in an antibody-dependent cell cytotoxicity (ADCC) reporter assay with FcγRIIIa-overexpressing T-cells. This could be explained with a stronger binding to the FcγRIIIa. Importantly, the novel construct could mediate a specific ADCC effect with natural killer cells similar to the parental antibody. |
| Databáze: | OpenAIRE |
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