Structural studies on 2’-O-methyltransferase TrmL that modifies the wobble nucleotide in tRNA_Leu

Abstract

The methyl-transfer reaction is one of the most universal and common RNA modifications. The methylated nucleoside is important for protein synthesis and stability of the tRNA structure. One of the methyltransferases, TrmL is responsible for 2'-O-methylation at the wobble position of the tRNALeu (CAA/UAA). Unlike the SPOUT methyltransferase superfamily, previously reported crystal structures of TrmL contain only the catalytic SPOUT domain lacking the extension domain for tRNA binding. TrmL from Escherichia coli (EcTrmL) requires i6A37 modification in tRNALeu. Meanwhile, tRNALeu from Gram-positive Mycoplasma capricolum contains m6A at the equivalent position instead, implying the substrate specificity of McTrmL must be different. Interestingly, our crystallographic study shows that McTrmL contains an extension domain in the C-terminus unlike homologs in Gram-negative bacteria. Additionally, we determined the crystal structures of TrmL from Gram-negative Vibrio vulnificus (VvTrmL) and Shewanella oneidensis (SoTrmL). The cocrystal structure of VvTrmL complexed with a reaction product S-adenosyl-l-homocysteine (SAH) and the ligand-free structure show that the binding of cofactor triggers conformational changes around the knot loops and flipping of E105, a putative general base required for activating the 2’-OH group of the modified nucleoside.