A chemical method for labeling lysine methyltransferase substrates.

Area of Science:

• Epigenetics
• Chemical Biology
• Proteomics

Background:

• Protein lysine methyltransferases (PKMTs) are crucial for regulating chromatin-dependent processes.
• Many PKMTs have uncharacterized substrates beyond histones, limiting our understanding of their functions.
• Existing methods for substrate identification are often biased or lack proteome-wide coverage.

Purpose of the Study:

• To develop an unbiased, chemical biology approach for identifying novel PKMT substrates.
• To enable proteome-wide discovery of non-histone PKMT targets.
• To provide a versatile tool for epigenetics research.

Main Methods:

• Utilized an alkyne-bearing S-adenosylmethionine (SAM) analogue as a cofactor for PKMTs.
• Employed the PKMT SETDB1 to enzymatically transfer the alkyne moiety to its substrates.
• Used click chemistry with azide-functionalized probes to label and affinity-purify PKMT substrates.

Main Results:

• Successfully demonstrated the proof-of-concept by labeling a recombinant histone H3 substrate with SETDB1.
• Developed a method for unbiased, proteome-wide identification of PKMT substrates.
• Showcased the potential for labeling and purifying multiple substrates simultaneously from complex biological extracts.

Conclusions:

• The described chemical biology strategy offers a powerful tool for unbiased PKMT substrate discovery.
• This method has broad applicability in epigenetics for identifying novel protein lysine methyltransferase targets.
• The approach facilitates simultaneous labeling and purification of numerous substrates, advancing proteomic analysis.