Nucleoside-modified AdoMet analogues for differential methyltransferase targeting

Area of Science:

• Biochemistry
• Chemical Biology
• Enzymology

Background:

• Methyltransferases (MTases) are crucial enzymes that utilize S-adenosyl-l-methionine (AdoMet) for methyl group transfer to various biomolecules.
• Synthetic AdoMet analogues offer potential for site-specific functionalization but often lack selectivity among different MTases.
• Current strategies, like modifying the sulfur/selenium substituent, are insufficient to differentiate between promiscuous MTases.

Purpose of the Study:

• To design and synthesize novel AdoMet analogues with improved selectivity for distinct methyltransferases.
• To investigate the impact of modifications on the nucleoside moiety of AdoMet on MTase activity and substrate discrimination.
• To establish a method for producing these novel analogues using a characterized methionine adenosyltransferase (MAT).

Main Methods:

• Synthesis of a panel of AdoMet analogues with variations in the nucleoside moiety (NM-AdoMets).
• Enzymatic production of NM-AdoMets using a previously uncharacterized methionine adenosyltransferase (MAT) with methionine and ATP analogues (e.g., ITP, N⁶-propargyl-ATP).
• Assay of NM-AdoMet activity with three representative MTases to determine substrate discrimination and relative activity changes.

Main Results:

• Efficient production of NM-AdoMets was achieved using a novel MAT enzyme.
• N⁶-modification of the AdoMet nucleoside moiety altered the relative activity of tested MTases by up to 13-fold.
• These modifications successfully discriminated between substrates for methyl transfer, demonstrating improved MTase selectivity.
• The N⁶-modified analogues could also facilitate the transfer of allyl and propargyl groups.

Conclusions:

• Novel NM-AdoMets provide a powerful tool for achieving enhanced selectivity among methyltransferases.
• Modification of the AdoMet nucleoside moiety offers a viable strategy to overcome limitations of existing AdoMet analogues in discriminating between promiscuous MTases.
• These engineered AdoMet analogues expand the toolkit for precise control of enzymatic methylation and functional group introduction in chemical biology.