Page 126 - Tyrosine-Based Bioconjugations - Jorick Bruins

Appendix
Introduction
Strain-promoted oxidation-controlled ortho-quinone cycloaddition (SPOCQ) is a reaction based on the facile (4+2) cycloaddition of an ortho-quinone (also known as 1,2-quinone) with a strained alkyne or strained alkene (Figure 1).1-3 Within the class, cyclopropanated versions of cyclooctyne (bicyclo[6.1.0]nonyne, BCN).2, 4 and trans-cyclooctene (cpTCO)3, 5 were found to undergo particularly fast cycloaddition, similar to accelerated cycloaddition with azido groups, by virtue of the additional strain imparted by the fused cyclopropane ring.
Figure 1. Concept of SPOCQ of an ortho-quinone with strained alkyne BCN or strained alkene cpTCO.
While of general interest to the field of metal-free click chemistry, a particularly useful aspect of an ortho-quinone is that it can be readily generated from a (substituted) phenol group by oxidation. Moreover, as the phenol moiety is present in the canonical amino acid tyrosine, in situ generation of an ortho-quinone moiety provides an excellent opportunity for application in the field of protein labelling. However, it is also apparent that global oxidation of tyrosine side-chains in a protein, for example by means of Fremy’s salt, will result in a heterogeneous product with varying amounts of conjugated label. One elegant approach to site-selective labelling entails the genetic encoding of L-DOPA (3,4-dihydroxyphenylalanine) followed by mild selective oxidation.6 However, genetic encoding of non-canonical amino acids often provides relatively modest protein titers.
A promising approach to directed protein labelling based on canonical amino acids involves the introduction of an exposed tyrosine residue in a C- or N-terminal peptide fusion, e.g. tetra- glycyltyrosine (G4Y), which enables the chemoselective oxidation of the exposed tyrosine moiety exclusively upon exposure to the enzyme mushroom tyrosinase (mTyr).7, 8 Thus, in the presence
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