A new strategy to identify the protein substrates of specific protein methyltransferases should allow scientists to better delineate the roles these signaling enzymes play in cells (J. Am. Chem. Soc., DOI: 10.1021/ja2118333). Methyltransferases, many of which are involved in gene transcription and epigenetic regulation, transfer a methyl group from an S-adenosyl-l-methionine (SAM) cofactor to an amino acid side chain on a protein substrate. To pinpoint the targets of a given methyltransferase, Minkui Luo and Kabirul Islam of Memorial Sloan-Kettering Cancer Center and coworkers tweak the enzyme’s active site so that it can accept a version of SAM that donates a 4-azido-but-2-enyl group rather than the usual methyl. When this azide-containing SAM is added to the contents of cells that express the altered methyltransferase, the enzyme transfers the azide group to its substrates. Luo’s team then uses copper-free click chemistry and mass spectrometry to isolate and identify the enzyme’s substrates. The method “will be helpful in understanding the full spectrum of potential intracellular substrates of specific protein methyltransferases,” comments Robert A. Copeland of Cambridge, Mass.-based Epizyme, which is developing small-molecule methyltransferase inhibitors. But the method’s scope remains to be tested, he adds, because the SAM-binding pockets of these enzymes have diverse structures.