Site-specific incorporation of phosphotyrosine using an expanded genetic code

Access to phosphoproteins with stoichiometric and site-specific phosphorylation status is key for understanding the role of protein phosphorylation. Here we report an efficient method to generate pure, active phosphotyrosine-containing proteins by genetically encoding a stable phosphotyrosine analogue that is convertible into native phosphotyrosine. We demonstrate its general compatibility with proteins of various sizes, phosphotyrosine site and function, and reveal a possible negative regulation role of tyrosine phosphorylation in ubiquitination.

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