A chemical method for labeling lysine methyltransferase substrates.
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Several protein lysine methyltransferases (PKMTs) modify histones to regulate chromatin-dependent cellular processes, such as transcription, DNA replication and DNA damage repair. PKMTs are likely to have many additional substrates in addition to histones, but relatively few nonhistone substrates have been characterized, and the substrate specificity for many PKMTs has yet to be defined. Thus, new unbiased methods are needed to find PKMT substrates. Here, we describe a chemical biology approach for unbiased, proteome-wide identification of novel PKMT substrates. Our strategy makes use of an alkyne-bearing S-adenosylmethionine (SAM) analogue, which is accepted by the PKMT, SETDB1, as a cofactor, resulting in the enzymatic attachment of a terminal alkyne to its substrate. Such labeled proteins can then be treated with azide-functionalized probes to ligate affinity handles or fluorophores to the PKMT substrates. As a proof-of-concept, we have used SETDB1 to transfer the alkyne moiety from the SAM analogue onto a recombinant histone H3 substrate. We anticipate that this chemical method will find broad use in epigenetics to enable unbiased searches for new PKMT substrates by using recombinant enzymes and unnatural SAM cofactors to label and purify many substrates simultaneously from complex organelle or cell extracts.
Staining and Labeling
Published Version (Please cite this version)10.1002/cbic.201000433
Publication InfoBinda, Olivier; Boyce, Michael; Rush, Jason S; Palaniappan, Krishnan K; Bertozzi, Carolyn R; & Gozani, Or (2011). A chemical method for labeling lysine methyltransferase substrates. Chembiochem : a European journal of chemical biology, 12(2). pp. 330-334. 10.1002/cbic.201000433. Retrieved from https://hdl.handle.net/10161/19701.
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Michael Scott Boyce
Associate Professor of Biochemistry
The Boyce Lab studies mammalian cell signaling through protein glycosylation. For the latest news, project information and publications from our group, please visit our web site at http://www.boycelab.org or follow us on Twitter at https://twitter.com/BoyceLab.
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