Fluorophore-Conjugated Holliday Junctions for Generating Super-Bright Antibodies and Antibody Fragments
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Ploegh_Fluorophore.pdf
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Author(s)
Theile, Christopher S.
Chen, Guan-Yu
Bilate, Angelina M.
Duarte, Joao N.
Avalos, Ana M.
Fang, Tao
Barberena, Roberto
Sato, Shuji
Ploegh, Hidde
Li, Zeyang,S.M.Massachusetts Institute of Technology.
Date Issued
September 2015
Journal
Angewandte Chemie International Edition
Publisher
Wiley Blackwell
Citation
Li, Zeyang et al. “Fluorophore-Conjugated Holliday Junctions for Generating Super-Bright Antibodies and Antibody Fragments.” Angewandte Chemie International Edition 54.40 (2015): 11706–11710.
Version
Author's final manuscript
Abstract
The site-specific modification of proteins with fluorophores can render a protein fluorescent without compromising its function. To avoid self-quenching from multiple fluorophores installed in close proximity, we used Holliday junctions to label proteins site-specifically. Holliday junctions enable modification with multiple fluorophores at reasonably precise spacing. We designed a Holliday junction with three of its four arms modified with a fluorophore of choice and the remaining arm equipped with a dibenzocyclooctyne substituent to render it reactive with an azide-modified fluorescent single-domain antibody fragment or an intact immunoglobulin produced in a sortase-catalyzed reaction. These fluorescent Holliday junctions improve fluorescence yields for both single-domain and full-sized antibodies without deleterious effects on antigen binding.
MIT Department
Massachusetts Institute of Technology. Department of Biology
Whitehead Institute for Biomedical Research
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Creative Commons Attribution-Noncommercial-Share Alike
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DOI of Published Version
https://doi.org/10.1002/anie.201505277
