Purification and properties of several transfer RNA methyltransferases fromS. typhimurium

Summary A fast method for a single-step fractionation of a number of tRNA methyltransferases fromSalmonella typhimurium is described. The method basically consists of ion-exchange chromatography on a phosphocellulose column and permits the separation of the enzymes forming mt⁶A, m¹G, m⁵U, m⁷G. The enzyme fractions appear sufficiently purified to allow the estimation of some molecular and kinetic properties. The apparent K_M for adenosylmethionine range between 1.5 to 3.2×10⁻⁵ M, whereas K_M for undermethylated tRNA range between 3.1×10⁻⁵ M to 3.1×10⁻⁴ M. Glycerol gradient determination indicates the following M_r for the native proteins: 25×10³, 40×10³, 50×10³ and 65×10³ for m⁷G-, mt⁶A-, m¹G- and m⁵U-forming enzymes, respectively. A complete analysis of methylated nucleosides formedin vivo inS. typhimurium has been obtained: it also allowed us to infer the pattern of the various tRNA methyltransferases for this prokaryote. The tRNA methyltransferase forming mt⁶A has been isolated for the first time from any type of cell.