Abstract
Within the realm of lysine methylation, the discovery of lysine methyltransferase (KMTs) substrates has been burgeoning because of established systematic substrate screening protocols. Here, we describe a protocol enabling the systematic identification of JmjC KDM substrate preference and in vitro substrates. Systematically designed peptide libraries containing methylated lysine residues are used to characterize enzyme-substrate preference and identify new candidate substrates in vitro. For complete details on the use and execution of this protocol, please refer to Hoekstra and Biggar (2021).