A Platform for Site‐Specific DNA‐Antibody Bioconjugation by Using Benzoylacrylic‐Labelled Oligonucleotides. (2nd November 2021)

• Record Type: Journal Article
• Authors: Konč, Juraj; Brown, Libby; Whiten, Daniel R.; Zuo, Yukun; Ravn, Peter; Klenerman, David; Bernardes, Gonçalo J. L.
• Is Part Of: Angewandte Chemie. Volume 133:Number 49(2021)
• Page Start: 26109
• Page End: 26117
• Publication Date: 2021-11-02
• Subjects: antibodies -- bioconjugation -- imaging -- mass spectrometry -- nucleic acids; Chemistry -- Periodicals; 540
• DOI: 10.1002/ange.202109713
• Languages: English
• ISSNs: 0044-8249

Abstract

Abstract: Many bioconjugation strategies for DNA oligonucleotides and antibodies suffer limitations, such as site‐specificity, stoichiometry and hydrolytic instability of the conjugates, which makes them unsuitable for biological applications. Here, we report a new platform for the preparation of DNA‐antibody bioconjugates with a simple benzoylacrylic acid pentafluorophenyl ester reagent. Benzoylacrylic‐labelled oligonucleotides prepared with this reagent can be site‐specifically conjugated to a range of proteins and antibodies through accessible cysteine residues. The homogeneity of the prepared DNA‐antibody bioconjugates was confirmed by a new LC‐MS protocol and the bioconjugate probes were used in fluorescence or super‐resolution microscopy cell imaging experiments. This work demonstrates the versatility and robustness of our bioconjugation protocol that gives site‐specific, well‐defined and plasma‐stable DNA‐antibody bioconjugates for biological applications. Abstract : BA‐PFP is a simple and practical reagent that enables preparation of a range of diverse site‐specific, homogenous DNA‐protein/antibody bioconjugates under mild conditions. The generated constructs can be conveniently characterized by an optimized LC‐MS method and utilized as probes for cell imaging using fluorescence or super‐resolution DNA‐PAINT microscopy.