Quaternization of Vinyl/Alkynyl Pyridine Enables Ultrafast Cysteine-Selective Protein Modification and Charge Modulation.
Quaternized vinyl- and alkynyl-pyridine reagents were shown to react in an ultrafast and selective manner with several cysteine-tagged proteins at near-stoichiometric quantities. We have demonstrated that this method can effectively create a homogenous antibody-drug conjugate that features a precise drug-to-antibody ratio of 2, which was stable in human plasma and retained its specificity towards Her2+ cells. Finally, the developed warhead introduces a +1 charge to the overall net charge of the protein, which enabled us to show that the electrophoretic mobility of the protein may be tuned through the simple attachment of a quaternized vinyl pyridinium reagent at the cysteine residues. We anticipate the generalized use of quaternized vinyl- and alkynyl-pyridine reagents not only for bioconjugation, but also as warheads for covalent inhibition and as tools to profile cysteine reactivity.
['Matos|Maria J|MJ|', 'Navo|Claudio D|CD|', 'Hakala|Tuuli|T|', 'Ferhati|Xhenti|X|', 'Guerreiro|Ana|A|', 'Hartmann|David|D|', 'Bernardim|Barbara|B|', 'Saar|Kadi L|KL|', 'Compañón|Ismael|I|', 'Corzana|Francisco|F|', 'Knowles|Tuomas P J|TPJ|', 'Jiménez-Osés|Gonzalo|G|', 'Bernardes|Gonçalo J L|GJL|0000-0001-6594-8917']
protein modification antibody-drug conjugates cysteine bioconjugation microfluidics