Characterization of a mimivirus RNA cap guanine-N2 methyltransferase

Methyl acceptor specificity of MimiTgs. Reaction mixtures (10 μL) containing 50 mM Tris-acetate (pH 6.0), 5 mM DTT, 50 μM [³H-CH₃]AdoMet, 1 mM methyl acceptor as specified, and 1 μg MimiTgs were incubated for 60 min at 37°C. Aliquots (4 μL) were spotted on PEI-cellulose TLC plates, which were developed either with 0.05 M (NH₄)₂SO₄ (for reactions with GpppA, m⁷GpppA, GpppG, and m⁷GpppG as methyl acceptors) or 0.1 M (NH₄)₂SO₄ (for reactions with GDP and m⁷GDP as methyl acceptors) or 0.2 M (NH₄)₂SO₄ (for reactions with GTP, and m⁷GTP as methyl acceptors). The chromatograms were treated with Enhance and ³H-labeled material was visualized by autoradiography. The methyltransferase reaction products m^2,7GpppA, m^2,7GpppG, m^2,7GDP, and m^2,7GTP are denoted by arrowheads at the right of the chromatograms.