|Title||Inducible, selective labeling of proteins via enzymatic oxidation of tyrosine|
|Author(s)||Bruins, Jorick J.; Wouw, Criss van de; Keijzer, Jordi F.; Albada, Bauke; Delft, Floris L. van|
|Source||In: Methods in Molecular Biology Hertfordshire : Springer (Methods in Molecular Biology ) - ISBN 9781493995455 - p. 357 - 368.|
|Publication type||Peer reviewed book chapter|
|Keyword(s)||Antibody–drug conjugates (ADC) - Bioconjugate chemistry - Bioorthogonal click - Protein functionalization - SPOCQ - Tyrosine modification|
Proteins can be labeled site-specifically and in inducible fashion by exposing a small peptide tag (G4Y) on any of its termini and activating the newly exposed tyrosine residue with the enzyme mushroom tyrosinase. The enzyme generates a quinone by oxidizing the tyrosine, which in turn can perform strain-promoted oxidation-controlled ortho-quinone cycloaddition (SPOCQ) with strained alkynes and alkenes, generating a stable conjugation product. Here, we describe a protocol to perform SPOCQ reaction on proteins, along with notes to optimize yield and reaction rates. Conjugation efficiencies of over 95% to antibodies have been reported using this protocol.