Recent development of methods for genetic incorporation of unnatural amino acids into proteins in live cells enables us to analyze protein interactions by site-specific photocrosslinking. Here we describe a method to incorporate p-benzoyl-L-phenylalanine (pBpa), a photoreactive unnatural amino acid, into defined positions of a target protein in living yeast cells. Photocrosslinking using the pBpa-incorporated proteins has been proven to be a powerful method for analyzing protein-protein interactions at the spatial resolution of amino-acid residues. Since photocrosslinking can be performed for pBpa-incorporated proteins that are properly assembled into a protein complex in living cells, this method will allow us to reveal protein-protein interactions of the target proteins at work.

• PMID: 23996180
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Shiota T, Nishikawa S, Endo T

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