Journal: JACS Au

Article Title: Aminothiazolone Inhibitors Disrupt the Protein–RNA Interaction of METTL16 and Modulate the m 6 A RNA Modification

doi: 10.1021/jacsau.3c00832

Figure Lengend Snippet: Aminothiazolones bind to METTL16 and inhibit the methyltransferase activity of METTL16. (A) Δ T m values of different inhibitors. Data are shown as mean ± SEM, n = 4. (B) Denaturation curves of METTL16 with tested compounds. In contrast to the control compound Ia , inhibitors 27 , 45 , 46 , and 47 , dose-dependently change the thermal stability of the METTL16 protein. Data are shown as mean ± SD, n = 2. (C) and (D) Biosensor assay evaluated the binding affinities of compounds 45 and 47 toward METTL16. (E) Schematic of aminothiazolones inhibiting the methyltransferase activity of METTL16 through disruption of the protein–RNA interaction of METTL16 and MAT2A- hp1 mRNA. Inhibitors were preincubated with the METTL16 MTD protein for 30 min, and a mixture of MAT2A -hp1 and SAM was added to the reaction. After reacting for 1 h at room temperature, MTase Glo reagents were added, and the luminescence was measured by TECAN. (F) Compounds 27 , 45 , 46 , and 47 dose-dependently inhibited the methyltransferase activity of METTL16. The potassium salt of compound 45 was used in the biosensor assay. Data are shown as mean ± SD, n = 2.

Article Snippet: The methyltransferase activity of METTL16 MTD was measured using an MTase Glo kit (Promega V7601) following the manufacturer’s instructions.

Techniques: Activity Assay, Control, Biosensor Assay, Binding Assay, Disruption

Journal: JACS Au

Article Title: Aminothiazolone Inhibitors Disrupt the Protein–RNA Interaction of METTL16 and Modulate the m 6 A RNA Modification

doi: 10.1021/jacsau.3c00832

Figure Lengend Snippet: Molecular docking analysis of compound 45 with METTL16. (A) The catalytic N -terminal methyltransferase domain of METTL16 in complex with MAT2A -hp1 3′UTR hairpin (PDB: 6DU4, residues 1–310). The protein is shown in the charged surface. (B) An optimal docking configuration of 45 in complex with the RNA-binding site of METTL16 (PDB: 6DU4 ). METTL16 is shown as the charged surface and 45 as the green carbon backbone. (C) The ribbon illustration of METTL16 (in cyan) with 45 (in green carbon backbone). Selected key interacting residues are depicted as sticks. (D) 2D illustration of the interaction between 45 and METTL16. Residues directly involved in compound interaction are indicated in blue circles, and residues indirectly involved in compound interaction are indicated in cyan circles. The proposed binding mode shows a salt bridge (Lys251) and a hydrogen bond (Asp198) between METTL16 and the carboxylic acid residue of 45 . Arg204 thereby forms additional hydrogen bonds with sulfonamide oxygens. Depicted in green, 45 forms a π–π interaction with Trp283.

Article Snippet: The methyltransferase activity of METTL16 MTD was measured using an MTase Glo kit (Promega V7601) following the manufacturer’s instructions.

Techniques: RNA Binding Assay, Binding Assay, Residue

Journal: bioRxiv

Article Title: A mitochondrial SET domain protein is essential in T. brucei and required for mitochondrial morphology and function

doi: 10.1101/2024.02.23.581798

Figure Lengend Snippet: A. Coomassie-stained gel and western blot of immunoprecipitations from 29-13 or 29-13 TbSETD.HA probed with anti-HA. HC; heavy chain LC; light chain B: Methyltransferase activity of control human SET7/9 and immunoprecipitations fractions from parental 29-13 and 29-13 TbSETD.HA. Two biological replicates are presented with each assay done in triplicate. RLU; relative luciferase activity.

Article Snippet: Assays were performed using Promega MTase-Glo Methyltransferase Assay (Promega V7601) with the following modifications.

Techniques: Staining, Western Blot, Activity Assay, Luciferase