Yeast cells expressing Su9-APEX2 are incubated with each probe at 2.5 mM final concentration for half an hour, and the labeling reaction is initiated by the addition of 1 mM H2O2. Alkyne tyramide is the most reactive APEX2 substrate for labeling the yeast proteome.
APEX2-mediated Alkyne tyramide labeling in the yeast mitochondria is more effective at probe concentrations above 1 mM.
Alkyne tyramide enables proteomic profiling in the mitochondrial matrix of intact yeast cells with exceptionally high specificity (94%), and offers higher coverage than the traditional APEX2 substrate biotin-phenol (BP).