Chemo- and Regioselective Lysine Modification on Native Proteins
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|Title:||Chemo- and Regioselective Lysine Modification on Native Proteins
|Author:||Matos, Maria João Correia Pinto Carvalho de
Oliveira, Bruno L.
Martínez Saéz, Nuria
Cal, Pedro M.S.D.
Bertoldo, Jean Borges
Maneiro Rey, María
Howard, Julie A.
Deery, Michael J.
Chalker, Justin M.
Jiménez Osés, Gonzalo
Bernardes, Gonçalo J.L.
|Affiliation:||Universidade de Santiago de Compostela. Centro de Investigación en Química Biolóxica e Materiais Moleculares
Universidade de Santiago de Compostela. Departamento de Química Orgánica
|Subject:||Reagents | Modification | Peptides and proteins | Monomers | Organic compounds ||
|Date of Issue:||2018
|Publisher:||American Chemical Society
|Citation:||J. Am. Chem. Soc. 2018, 140, 11, 4004–4017
|Abstract:||Site-selective chemical conjugation of synthetic molecules to proteins expands their functional and therapeutic capacity. Current protein modification methods, based on synthetic and biochemical technologies, can achieve site selectivity, but these techniques often require extensive sequence engineering or are restricted to the N- or C-terminus. Here we show the computer-assisted design of sulfonyl acrylate reagents for the modification of a single lysine residue on native protein sequences. This feature of the designed sulfonyl acrylates, together with the innate and subtle reactivity differences conferred by the unique local microenvironment surrounding each lysine, contribute to the observed regioselectivity of the reaction. Moreover, this site selectivity was predicted computationally, where the lysine with the lowest pKa was the kinetically favored residue at slightly basic pH. Chemoselectivity was also observed as the reagent reacted preferentially at lysine, even in those cases when other nucleophilic residues such as cysteine were present. The reaction is fast and proceeds using a single molar equivalent of the sulfonyl acrylate reagent under biocompatible conditions (37 °C, pH 8.0). This technology was demonstrated by the quantitative and irreversible modification of five different proteins including the clinically used therapeutic antibody Trastuzumab without prior sequence engineering. Importantly, their native secondary structure and functionality is retained after the modification. This regioselective lysine modification method allows for further bioconjugation through aza-Michael addition to the acrylate electrophile that is generated by spontaneous elimination of methanesulfinic acid upon lysine labeling. We showed that a protein–antibody conjugate bearing a site-specifically installed fluorophore at lysine could be used for selective imaging of apoptotic cells and detection of Her2+ cells, respectively. This simple, robust method does not require genetic engineering and may be generally used for accessing diverse, well-defined protein conjugates for basic biology and therapeutic studies|
|Rights:||Copyright © 2018 American Chemical Society. This is an open access article published under a Creative Commons Attribution (CC-BY) License, which permits unrestricted use, distribution and reproduction in any medium, provided the author and source are cited
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Except where otherwise noted, this item's license is described as Copyright © 2018 American Chemical Society. This is an open access article published under a Creative Commons Attribution (CC-BY) License, which permits unrestricted use, distribution and reproduction in any medium, provided the author and source are cited