Proteomics

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RPL40 methyltransferase discovery and characterisation

ABSTRACT: The eukaryotic ribosome is highly modified by protein methylation, yet many of the responsible methyltransferases remain unknown. Here we have identified SMYD5 as a ribosomal protein methyltransferase that catalyses trimethylation of RPL40 at lysine 22. Through a systematic mass spectrometry-based approach, we show that the human ribosome has 12 primary sites of protein methylation, including at RPL40 K22. Through in vitro methylation of synthetic RPL40 using fractionated lysate, we then identify SMYD5 as a candidate RPL40 K22 methyltransferase. We show that recombinant SMYD5 has robust activity towards RPL40 K22 in vitro, and that active site mutations ablate this activity. Knockouts of SMYD5 in K562 cells show a complete loss of RPL40 K22 methylation and decrease polysome levels. By systematic analysis of its recognition motif, we show that SMYD5 requires a tyrosine in the +2 position, and thereby is incapable of methylating its previously reported histone substrates.

ORGANISM(S): Homo Sapiens

SUBMITTER: Joshua Hamey  

PROVIDER: PXD056489 | panorama | Wed Apr 16 00:00:00 BST 2025

REPOSITORIES: PanoramaPublic

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SMYD5 is a ribosomal methyltransferase that trimethylates RPL40 lysine 22 through recognition of a KXY motif.

Hamey Joshua J JJ   Shah Manan M   Wade John D JD   Bartolec Tara K TK   Wettenhall Richard E H REH   Quinlan Kate G R KGR   Williamson Nicholas A NA   Wilkins Marc R MR  

Cell reports 20250405 4

The eukaryotic ribosome is highly modified by protein methylation, yet many of the responsible methyltransferases remain unknown. Here, we identify SET and MYND domain-containing protein 5 (SMYD5) as a ribosomal protein methyltransferase that catalyzes trimethylation of RPL40/eL40 at lysine 22. Through a systematic mass spectrometry-based approach, we identify 12 primary sites of protein methylation in ribosomes from K562 cells, including at RPL40 K22. Through in vitro methylation of synthetic R  ...[more]

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