N2-Guanine specific transfer RNA methyltransferase I from rat liver and leukemic rat spleen

Abstract

An enzyme was purified from rat liver and leukemic rat spleen wnich methylates guanosine residues in tRNA to li²-methylguanosine. By sequence analysis of bulk E. coli tRNA methylated with crude extracts it was shown that the enzyme is responsible for about 50% of total m²G formed in vitro. The extent of methylation of a number of homogenous tRNA species was measured using the purified enzyme from both sources. Among tested E. coli tRNAs only tRNA^Arg, tRNA^Phe, and tRNA^Val yielded significantly more m²G than the bulk tRNA. The K_m for tRNA^Arg in the methylation reaction with enzymes from either tissue was 7.8 × 10^-7 M as compared to the value 1 × 10^-5 M obtained for the bulk tRNA. In a pancreatic RNase digest of bulk tRNA as well as of pure tRNA^Arg, tRNA^Phe, and tRNA^Val, A-m²G-Cp was found to be the only sequence methylated. Thus, the mammalian methyl-transferase specifically recognizes the guanylate residue at position 10 from the 5′-end contained in a sequence (s⁴)U-A-G-Cp. Furthermore, there is no change between the enzyme from normal liver and leukemic spleen in the affinity for tRNA, the methylating capacity, and tRNA site and sequence recognition specificity.