Specificity analysis of human protein lysine methyltransferases and proteomwide identification of novel substrate proteins

• Applicant: Professor Dr. Albert Jeltsch
• Subject Area: Biochemistry
• Term: from 2008 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 62953237

Protein lysine methyltransferases (PKMT) were discovered in 2000 when it was shown that SUV39H1 methylates histone H3 at lysine 9. However, several PKMTs that were initially identified as histone modifying enzymes were later found to methylate non-histone substrates as well and protein lysine methylation has been recognized as an important post-translational modification involved in various processes. Today, more than 8000 lysine methylation events are reported in the human proteome (Phosphosite Plus, 2016), but due to limitations in proteomics approaches, it is likely that many more are still to be discovered. Moreover, for most described lysine methylation events the responsible PKMT has not been identified and for many of them, the biological role of the methylation is not known. Moreover, for most PKMTs only few substrates have been identified up to now. For this reason, the identification of enzymes responsible for specific protein methylation events is a critical challenge for biochemical research. Moreover, sophisticated experiments addressing the cellular role of specific lysine methylation events are urgently needed. We developed and successfully employed a novel approach for the identification of non-histone targets of PKMTs. It starts with the determination of the substrate specificity of PKMTs with peptide arrays. The specificity profile can then be used to search the human proteome for candidate substrates. The methylation of these peptide and protein substrates is then investigated in vitro and in cells. Finally, we plan to develop cellular assays to uncover the biological role of the methylation event. The goals of this application are to continue this work and further advance it as described in the following work packages (WP):WP1: Cloning, expression and purification of additional human PKMTsWP2: Determination of the specificity profile of PKMTs and identify novel peptide substratesWP3: Identification of novel protein substrates in vitro and in cellsWP4: Investigation of the cellular role of the methylation eventsWP5: Development of a Web server allowing to identify pairs of PKMTs and lysine methylation events to make the results of our study available for all researches in the field.

• DFG Programme: Research Grants