Analysis of GTP addition in the reverse (3′–5′) direction by human tRNA^His guanylyltransferase

Structural comparison of Thg1 and TLP enzymes. (A) Human mitochondrial tRNA^His (hmtRNA^His) is drawn as a cloverleaf structure. hmtRNA^His contains a His-specific anticodon GUG and cytosine opposite the −1 position (Y = C). (B) Structural comparison of Thg1 enzymes: HsThg1 in the presence of hmtRNA^His (HsThg1-tetramer, this study), HsThg1-dimer (PDB ID: 3OTC) and CaThg1-free (PDB ID: 3WBZ), without tRNA^His binding. The flexibility of the β-hairpin structure at the carboxy-terminal of Thg1 is shown in red. (C) Structural comparison of Thg1 enzymes with tRNA binding: HsThg1 in the presence of hmtRNA^His (HsThg1-tetramer, this study), CaThg1-tRNA^His (PDB ID: 3WC2), and MaTLP-tRNA (PDB ID: 5AXN). The unknown electron density map (2Fo-Fc map [1.5 σ]; blue, and Fo-Fc map [2.5 σ]; green) close to the flexible β-hairpin is indicated as mesh. (D) Proposed incoming nucleotide recognition mechanism of HsThg1 organized by the interaction between the β-hairpin and the elbow region of the substrate tRNA.