Site-selective modification of antibodies using highly reactive chemical species

Abstract

We have developed protein modification reactions using highly reactive chemical species. Our focus includes the utilization of the highly reactive singlet oxygen (¹O₂) to selectively modify histidine residues (His). By constructing a reaction field on magnetic beads, bringing together the photocatalyst, and the Fc region of antibodies, we successfully functionalized His residues in the Fc domain. Furthermore, we harnessed highly reactive radical species to devise a novel reaction for selectively modifying tyrosine residues (Tyr). Leveraging the unique characteristics of Tyr residues in the IgG antibody structure, we developed a method to selectively functionalize the complementarity-determining regions. Evaluation of the selectivity of labeled sites was performed through papain digestion and gel electrophoresis. Expanding the applications, we employed the developed CDR-functionalized antibodies in creating fluorescent immunosensor molecules. The simplicity and efficiency of our chemical modification approach enabled the rapid transformation of therapeutic antibodies into fluorescent immunosensors.