Purification and characterization of tRNA (adenine-1-)-methyltransferase from Thermus flavus strain 71.

tRNA (adenine-1-)-methyltransferase was isolated from the extreme thermophile Thermus flavus, strain 71. It was purified about 2000-fold by ammonium sulfate fractionation and affinity chromatography on tRNA bound to aminohydroxybutylcellulose via its oxidized 3' end. The purified protein preparation is free of nuclease and aminoacyl-tRNA synthetase activity and contains no more than 4% of tRNA (guanine-7-)methyltransferase activity. The only activity of the enzyme is to methylate A58 in the T psi loop of tRNA. Out of the eight purified tRNAs examined, only yeast tRNATrp was not utilized as a substrate. The enzyme is highly thermostable. It is most active at 75 degrees C. tRNA (adenine-1-)-methyltransferase has a Km of 0.4-0.5 microM for tRNA2Gln from Escherichia coli and a Km of 6 microM for S-adenosyl-L-methionine.