The Role of Trp79 in ß-Actin on Histidine Methyltransferase SETD3 Catalysis.

Nτ -methylation of His73 in actin by histidine methyltransferase SETD3 plays an important role in stabilising actin filaments in eukaryotes. Mutations in actin and overexpression of SETD3 have been related to human diseases, including cancer. Here, we investigated the importance of Trp79 in ß-actin on productive human SETD3 catalysis. Substitution of Trp79 in ß-actin peptides by its chemically diverse analogues reveals that the hydrophobic Trp79 binding pocket modulates the catalytic activity of SETD3, and that retaining a bulky and hydrophobic amino acid at position 79 is important for efficient His73 methylation by SETD3. Molecular dynamics simulations show that the Trp79 binding pocket of SETD3 is ideally shaped to accommodate large and hydrophobic Trp79, contributing to the favourable release of water molecules upon binding. Our results demonstrate that the distant Trp79 binding site plays an important role in efficient SETD3 catalysis, contributing to the identification of new SETD3 substrates and the development of chemical probes targeting the biomedically important SETD3.

Importance of Ile71 in ß-actin on histidine methyltransferase SETD3 catalysis.

SETD3-catalysed N3-methylation of His73 in ß-actin plays a key role in stabilisation of actin filaments in the metazoan cells. Overexpression and/or dysregulation of SETD3 is associated with several human pathologies, including cancer. Here, we examined the role of the Ile71 residue in ß-actin on human SETD3 catalysis. Substitution of Ile71 in ß-actin peptides by its natural and unnatural mimics reveals that the 'secondary' Ile71 binding pocket modulates the substrate efficiency of ß-actin. Our enzymatic work demonstrates that human SETD3 can accommodate structurally diverse hydrophobic side chains in its Ile71 binding pocket, providing clear limits of the size and shape of Ile analogues. Water thermodynamics calculations reveal that the Ile71 pocket is occupied by high-energy water molecules, that are released upon the Ile71 binding, contributing favourably to the SETD3-ßA complex formation. The work highlights that the hydrophobic Ile71 binding site plays an essential role in SETD3 catalysis, contributing to an ongoing effort in the design and development of chemical probes targeting SETD3.

The Structure, Activity, and Function of the SETD3 Protein Histidine Methyltransferase.

SETD3 has been recently identified as a long sought, actin specific histidine methyltransferase that catalyzes the Nτ-methylation reaction of histidine 73 (H73) residue in human actin or its equivalent in other metazoans. Its homologs are widespread among multicellular eukaryotes and expressed in most mammalian tissues. SETD3 consists of a catalytic SET domain responsible for transferring the methyl group from S-adenosyl-L-methionine (AdoMet) to a protein substrate and a RuBisCO LSMT domain that recognizes and binds the methyl-accepting protein(s). The enzyme was initially identified as a methyltransferase that catalyzes the modification of histone H3 at K4 and K36 residues, but later studies revealed that the only bona fide substrate of SETD3 is H73, in the actin protein. The methylation of actin at H73 contributes to maintaining cytoskeleton integrity, which remains the only well characterized biological effect of SETD3. However, the discovery of numerous novel methyltransferase interactors suggests that SETD3 may regulate various biological processes, including cell cycle and apoptosis, carcinogenesis, response to hypoxic conditions, and enterovirus pathogenesis. This review summarizes the current advances in research on the SETD3 protein, its biological importance, and role in various diseases.