Site-Specific Bioconjugation through Enzyme-Catalyzed Tyrosine–Cysteine Bond Formation

• Published in: ACS central science Vol. 6; no. 9; pp. 1564 - 1571
• Main Authors: Lobba, Marco J, Fellmann, Christof, Marmelstein, Alan M, Maza, Johnathan C, Kissman, Elijah N, Robinson, Stephanie A, Staahl, Brett T, Urnes, Cole, Lew, Rachel J, Mogilevsky, Casey S, Doudna, Jennifer A, Francis, Matthew B
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 23.09.2020
• ISSN: 2374-7943; 2374-7951
• DOI: 10.1021/acscentsci.0c00940

The synthesis of protein–protein and protein–peptide conjugates is an important capability for producing vaccines, immunotherapeutics, and targeted delivery agents. Herein we show that the enzyme tyrosinase is capable of oxidizing exposed tyrosine residues into o-quinones that react rapidly with cysteine residues on target proteins. This coupling reaction occurs under mild aerobic conditions and has the rare ability to join full-size proteins in under 2 h. The utility of the approach is demonstrated for the attachment of cationic peptides to enhance the cellular delivery of CRISPR-Cas9 20-fold and for the coupling of reporter proteins to a cancer-targeting antibody fragment without loss of its cell-specific binding ability. The broad applicability of this technique provides a new building block approach for the synthesis of protein chimeras.