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Cited by 21Monoclonal and Polyclonal Antibodies ResearchReceptor Mechanisms and SignalingBiochemical and Structural Characterization

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Characterization of a Nanobody-Epitope Tag Interaction and Its Application for Receptor Engineering

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Cabalteja, Chino C., Sachdev, Shivani, Cheloha, Ross W.·2022·ACS Chemical Biology
DOI 10.1021/acschembio.2c00407PMC PMC10200313

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Abstract

Peptide epitope tags offer a valuable means for detection and manipulation of protein targets for which high quality detection reagents are not available. Most commonly used epitope tags are bound by conventional, full-size antibodies (Abs). The complex architecture of Abs complicates their application in protein engineering and intracellular applications. To address these shortcomings, single domain antibodies (nanobodies, Nbs) that recognize short peptide epitopes have become increasingly prized. Here, we characterize the interaction between a Nb (Nb6E) and a 14-mer peptide epitope. We identify residues in the peptide epitope essential for high affinity binding. Using this information in combination with computational modeling we propose a mode of interaction between Nb6E and this epitope. We apply this nanobody–epitope pair to augment the potency of a ligand at an engineered adenosine A2A receptor. This characterization of the nanobody–epitope pair opens the door to diverse applications including mechanistic studies of the G protein-coupled receptor function.

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Research works printed but not linked: 37

  1. Carter, 2018, Next Generation Antibody Drugs: Pursuit of the “High-Hanging Fruit, Nat. Rev. Drug Discovery, 17, 197, 10.1038/nrd.2017.227 · DOI
  2. Hutchings, 2020, A Review of Antibody-Based Therapeutics Targeting G Protein-Coupled Receptors: An Update, Expert Opin. Biol. Ther., 20, 925, 10.1080/14712598.2020.1745770 · DOI
  3. Vandemoortele, 2019, Pick a Tag and Explore the Functions of Your Pet Protein, Trends Biotechnol., 37, 1078, 10.1016/j.tibtech.2019.03.016 · DOI
  4. Chodorge, 2018, Engineering of a GLP-1 Analogue Peptide/Anti-PCSK9 Antibody Fusion for Type 2 Diabetes Treatment, Sci. Rep., 8, 17545, 10.1038/s41598-018-35869-4 · DOI
  5. Nelson, 2010, Antibody Fragments: Hope and Hype, mAbs, 2, 77, 10.4161/mabs.2.1.10786 · DOI
  6. Fujiwara, 2020, Impact of ScFv Structure in Chimeric Antigen Receptor on Receptor Expression Efficiency and Antigen Recognition Properties, Biochem. Biophys. Res. Commun., 527, 350, 10.1016/j.bbrc.2020.03.071 · DOI
  7. Cheloha, 2020, Exploring Cellular Biochemistry with Nanobodies, J. Biol. Chem., 295, 15307, 10.1074/jbc.REV120.012960 · DOI
  8. Boersma, 2019, Multi-Color Single-Molecule Imaging Uncovers Extensive Heterogeneity in MRNA Decoding, Cell, 178, 458, 10.1016/j.cell.2019.05.001 · DOI
  9. Traenkle, 2020, A Novel Epitope Tagging System to Visualize and Monitor Antigens in Live Cells with Chromobodies, Sci. Rep., 10, 14267, 10.1038/s41598-020-71091-x · DOI
  10. Götzke, 2019, The ALFA-Tag Is a Highly Versatile Tool for Nanobody-Based Bioscience Applications, Nat. Commun., 10, 4403, 10.1038/s41467-019-12301-7 · DOI
  11. Jedlitzke, 2022, A Light-Activatable Photocaged Variant of the Ultra-High Affinity ALFA-Tag Nanobody, ChemBioChem., 10.1002/cbic.202200079 · DOI
  12. De Genst, 2010, Structure and Properties of a Complex of α-Synuclein and a Single-Domain Camelid Antibody, J. Mol. Biol., 402, 326, 10.1016/j.jmb.2010.07.001 · DOI
  13. Braun, 2016, Peptides in Headlock--a Novel High-Affinity and Versatile Peptide-Binding Nanobody for Proteomics and Microscopy, Sci. Rep., 6, 19211, 10.1038/srep19211 · DOI
  14. Bradley, 2015, Potent and Efficacious Inhibition of CXCR2 Signaling by Biparatopic Nanobodies Combining Two Distinct Modes of Action, Mol. Pharmacol., 87, 251, 10.1124/mol.114.094821 · DOI
  15. Xu, 2022, Protein Visualization and Manipulation in Drosophila through the Use of Epitope Tags Recognized by Nanobodies, eLife, 11, e74326, 10.7554/eLife.74326 · DOI
  16. Ling, 2019, A Nanobody That Recognizes a 14-Residue Peptide Epitope in the E2 Ubiquitin-Conjugating Enzyme UBC6e Modulates Its Activity, Mol. Immunol., 114, 513, 10.1016/j.molimm.2019.08.008 · DOI
  17. Cheloha, 2020, Improved GPCR Ligands from Nanobody Tethering, Nat. Commun., 11, 2087, 10.1038/s41467-020-15884-8 · DOI
  18. Cheloha, 2021, Activation of a G Protein-Coupled Receptor through Indirect Antibody-Mediated Tethering of Ligands, RSC Chem. Biol., 2, 1692, 10.1039/D1CB00118C · DOI
  19. Pishesha, 2018, Sortase A: A Model for Transpeptidation and Its Biological Applications, Annu. Rev. Cell Dev. Biol., 34, 163, 10.1146/annurev-cellbio-100617-062527 · DOI
  20. Yang, 2020, Improved Protein Structure Prediction Using Predicted Interresidue Orientations, Proc. Natl. Acad. Sci. U. S. A., 117, 1496, 10.1073/pnas.1914677117 · DOI · Paper page
  21. Karle, 2001, Controls Exerted by the Aib Residue: Helix Formation and Helix Reversal This Article Is a US Government Work and, as Such, Is in the Public Domain in the United States of America, Biopolymers, 60, 351, 10.1002/1097-0282(2001)60:5<351::AID-BIP10174>3.0.CO;2-U · DOI
  22. Urban, 2015, DREADDs (Designer Receptors Exclusively Activated by Designer Drugs): Chemogenetic Tools with Therapeutic Utility, Annu. Rev. Pharmacol. Toxicol., 55, 399, 10.1146/annurev-pharmtox-010814-124803 · DOI
  23. Alvarez-Curto, 2015, Defining the Functional Equivalence of Wild-Type and Chemically Engineered G Protein-Coupled Receptors, 1, 10.1007/978-1-4939-2944-3_1 · DOI
  24. Irannejad, 2017, Functional Selectivity of GPCR-Directed Drug Action through Location Bias, Nat. Chem. Biol., 13, 799, 10.1038/nchembio.2389 · DOI
  25. Das, 2010, Nucleoside Conjugates of Quantum Dots for Characterization of G Protein-Coupled Receptors: Strategies for Immobilizing A2A Adenosine Receptor Agonists, J. Nanobiotechnology, 8, 11, 10.1186/1477-3155-8-11 · DOI
  26. Lebon, 2015, Molecular Determinants of CGS21680 Binding to the Human Adenosine A 2A Receptor, Mol. Pharmacol., 87, 907, 10.1124/mol.114.097360 · DOI
  27. Hong, 2009, Analysis and Optimization of Copper-Catalyzed Azide-Alkyne Cycloaddition for Bioconjugation, Angew. Chem., Int. Ed. Engl., 48, 9879, 10.1002/anie.200905087 · DOI
  28. Binkowski, 2011, A Luminescent Biosensor with Increased Dynamic Range for Intracellular CAMP, ACS Chem. Biol., 6, 1193, 10.1021/cb200248h · DOI
  29. Ramirez, 2020, Engineering a Proximity-Directed O-GlcNAc Transferase for Selective Protein O-GlcNAcylation in Cells, ACS Chem. Biol., 15, 1059, 10.1021/acschembio.0c00074 · DOI
  30. Mossner, 2020, Multimerization Strategies for Efficient Production and Purification of Highly Active Synthetic Cytokine Receptor Ligands, PLoS One, 15, e0230804, 10.1371/journal.pone.0230804 · DOI
  31. Fernandes, 2020, Immune Receptor Inhibition through Enforced Phosphatase Recruitment, Nature, 586, 779, 10.1038/s41586-020-2851-2 · DOI
  32. Xie, 2019, Nanobody-Based CAR T Cells That Target the Tumor Microenvironment Inhibit the Growth of Solid Tumors in Immunocompetent Mice, Proc. Natl. Acad. Sci. U. S. A., 116, 7624, 10.1073/pnas.1817147116 · DOI
  33. Mo, 2021, Nanobody-Based Chimeric Antigen Receptor T Cells Designed by CRISPR/Cas9 Technology for Solid Tumor Immunotherapy, Signal Transduct. Target. Ther., 6, 80, 10.1038/s41392-021-00462-1 · DOI
  34. Sachdev, 2021, Strategies for Targeting Cell Surface Proteins Using Multivalent Conjugates and Chemical Biology, Methods Cell Biol., 166, 205, 10.1016/bs.mcb.2021.06.004 · DOI
  35. Laeremans, 2022, Accelerating GPCR Drug Discovery With Conformation-Stabilizing VHHs, Front. Mol. Biosci., 9, 863099, 10.3389/fmolb.2022.863099 · DOI
  36. Cheloha, 2019, Internalization of Influenza Virus and Cell Surface Proteins Monitored by Site-Specific Conjugation of Protease-Sensitive Probes, ACS Chem. Biol., 14, 1836, 10.1021/acschembio.9b00493 · DOI
  37. Mirdita, 2022, ColabFold: Making Protein Folding Accessible to All, Nat. Methods, 19, 679, 10.1038/s41592-022-01488-1 · DOI · Paper page

Metadata

Authors
Cabalteja, C. C., Sachdev, S., & Cheloha, R. W.
Affiliations
National Institutes of Health (US)
Corresponding author
Cheloha, Ross W.
ORCID
Venue
ACS Chemical Biology
Publisher
American Chemical Society (ACS)
Published
2022-08-05
Volume
17
Issue
8
Pages
2296-2303
ISSN
  • 1554-8929 (ISSN-L)
  • 1554-8937
DOI
10.1021/acschembio.2c00407
PMC
PMC10200313
OpenAlex
W4289953209
Cited by
21
References
2
Citations by year
2026: 2 · 2025: 7 · 2024: 5 · 2023: 5 · 2022: 2
FWCI
2.70
Type
journal article
Language
en
Open access
Licence
Locations
  • ACS Chemical Biology (publishedVersion)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (acceptedVersion, public-domain)
Funders
  • National Institute of Diabetes and Digestive and Kidney Diseases
  • National Institutes of Health
Topics
  • Life Sciences › Immunology and Microbiology › Immunology › Monoclonal and Polyclonal Antibodies Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Receptor Mechanisms and Signaling
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Structural Characterization
Keywords
nanobodies, receptor engineering, adenosine A2A receptor, binding affinity, G protein-coupled receptors
MeSH terms
Antibodies, Epitopes, Peptides, Protein Engineering, Single-Domain Antibodies

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