Next, the capacity of EBX 2a to label tyrosine on proteins was assessed (Scheme 3). When the small regulatory native protein Ubiquitin (8.6 kDa) was submitted to the reaction conditions, nearly no reaction was observed. This confirmed that only accessible tyrosine can be functionalized and no other residues react. However, with denaturing conditions to gain access to the buried tyrosine together with a higher excess of EBX 2a (10 equivalents) and a longer reaction time (72 hours) (conditions A a ), modified ubiquitin 6 was obtained in 24% yield. Selective modification of residue Y59 was confirmed by targeted top-down mass spectrometry analysis (see Supporting Information for details). Higher conversion could be obtained using 50 equivalents of EBX 2a, but without increase in yield of 6 due to side reactions (conditions B a ). The same conditions A a were then applied to the hemeprotein myoglobin (17 kDa) containing 2 tyrosine residues. Chemo-and site-selectivity was achieved on the more reactive tyrosine Y146 to give 7 in 19% yield. Finally, we studied the functionalization of the tetrameric recombinant protein streptavidin (52 kDa), which contains 6 tyrosine residues per monomer (13 kDa), which are more accessible. Under native conditions (conditions A), 10 equivalents of EBX 2a allowed to mono-label streptavidin on the most accessible tyrosine Y31, Y42, Y48 or Y71 to give 8. This result reflects the reactivity and accessibility of these amino acids compared to Y10 and Y84, which were not modified.