ArticlemAbs
DNA immunization combined with scFv phage display identifies antagonistic GCGR specific antibodies and reveals new epitopes on the small extracellular loops.
Article in mAbs. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 19 papers.
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Who cites it
19 citing papers in PubMed, 33 citations in OpenAlex.
- Llama mRNA-LNP immunization enables rapid isolation of target specific nanobodies using cell-based panning.Antibody therapeutics · 2026Article
- TR-FRET between engineered nanobodies reveals the existence of endogenous CXCR4 oligomers.Communications biology · 2025Article
- Constitutive activity of an atypical chemokine receptor revealed by inverse agonistic nanobodies.Nature communications · 2025Article
- Structures of native SV2A reveal the binding mode for tetanus neurotoxin and anti-epileptic racetams.Nature communications · 2025Article
- Development of an anti-rat complement C2 antibody that improves renal outcome in a rat kidney transplant model.Frontiers in immunology · 2025Article
- Constitutive activity of an atypical chemokine receptor revealed by inverse agonistic nanobodies.bioRxiv : the preprint server for biology · 2024Article
- Discovery of Therapeutic Antibodies Targeting Complex Multi-Spanning Membrane Proteins.BioDrugs : clinical immunotherapeutics, biopharmaceuticals and gene therapy · 2024Review
- Structure elucidation of a human melanocortin-4 receptor specific orthosteric nanobody agonist.Nature communications · 2024Article
- Methods for Engineering Binders to Multi-Pass Membrane Proteins.Bioengineering (Basel, Switzerland) · 2023Review
- Revisiting the role of glucagon in health, diabetes mellitus and other metabolic diseases.Nature reviews. Endocrinology · 2023Review
- Conformational selection guides β-arrestin recruitment at a biased G protein-coupled receptor.Science (New York, N.Y.) · 2022Article
- Improving antibody drug development using bionanotechnology.Current opinion in biotechnology · 2022Review
- Antigen Design for Successful Isolation of Highly Challenging Therapeutic Anti-GPCR Antibodies.International journal of molecular sciences · 2020Review
- Phage Display Libraries for Antibody Therapeutic Discovery and Development.Antibodies (Basel, Switzerland) · 2019Review
- Glucagon Receptor Signaling and Lipid Metabolism.Frontiers in physiology · 2019Article
- Opportunities for therapeutic antibodies directed at G-protein-coupled receptors.Nature reviews. Drug discovery · 2017Review
- A cDNA Immunization Strategy to Generate Nanobodies against Membrane Proteins in Native Conformation.Frontiers in immunology · 2017Article
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Authors and funding
11 authors at 5 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The identification of functional monoclonal antibodies directed against G-protein coupled receptors (GPCRs) is challenging because of the membrane-embedded topology of these molecules. Here, we report the successful combination of llama DNA immunization with scFv-phage display and selections using virus-like particles (VLP) and the recombinant extracellular domain of the GPCR glucagon receptor (GCGR), resulting in glucagon receptor-specific antagonistic antibodies. By immunizing outbred llamas with plasmid DNA containing the human GCGR gene, we sought to provoke their immune system, which generated a high IgG1 response. Phage selections on VLPs allowed the identification of mAbs against the extracellular loop regions (ECL) of GCGR, in addition to multiple VH families interacting with the extracellular domain (ECD) of GCGR. Identifying mAbs binding to the ECL regions of GCGR is challenging because the large ECD covers the small ECLs in the energetically most favorable 'closed conformation' of GCGR. Comparison of Fab with scFv-phage display demonstrated that the multivalent nature of scFv display is essential for the identification of GCGR specific clones by selections on VLPs because of avid interaction. Ten different VH families that bound 5 different epitopes on the ECD of GCGR were derived from only 2 DNA-immunized llamas. Seven VH families demonstrated interference with glucagon-mediated cAMP increase. This combination of technologies proved applicable in identifying multiple functional binders in the class B GPCR context, suggesting it is a robust approach for tackling difficult membrane proteins.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.