Fast and Efficient Fc-Specific Photoaffinity Labeling To Produce Antibody-DNA Conjugates.

Stiller C, Aghelpasand H, Frick T, Westerlund K, Ahmadian A, Karlström AE

Bioconjug. Chem. 30 (11) 2790-2798 [2019-11-20; online 2019-10-25]

Antibody-DNA conjugates are powerful tools for DNA-assisted protein analysis. Growing usage of these methods demands efficient production of high-quality conjugates. We developed an easy and fast synthesis route yielding covalent antibody-DNA conjugates with a defined conjugation site and low batch-to-batch variability. We utilize the Z domain from protein A, containing the unnatural amino acid 4-benzoylphenylalanine (BPA) for photoaffinity labeling of the antibodies' Fc region. Z( xBPA) domains are C-terminally modified with triple-glycine (G3)-modified DNA-oligonucleotides via enzymatic Sortase A coupling. We show reliable modification of the most commonly used IgG's. To prove our conjugates' functionality, we detected antibody-antigen binding events in an assay called Droplet Barcode Sequencing for Protein analysis (DBS-Pro). It confirms not only retained functionality for both conjugate parts but also the potential of using DBS-Pro for quantifying protein abundances. As intermediates are easily storable and our approach is modular, it offers a convenient strategy for screening various antibody-DNA conjugates using the same starting material.

Bioinformatics Support for Computational Resources [Service]

NGI Stockholm (Genomics Applications) [Service]

NGI Stockholm (Genomics Production) [Service]

National Genomics Infrastructure [Service]

PubMed 31609586

DOI 10.1021/acs.bioconjchem.9b00548

Crossref 10.1021/acs.bioconjchem.9b00548

Publications 9.5.0