Abstract.-Transfer RNA fractions from Saccharomyces lactis and Saccharomyces cerevisiae have been tested for cytokinin activity in the tobacco bioassay. Cysteine tRNA has been identified as a cytokinin-containing tRNA species in S. cerevisiae. Acid hydrolysates of S. lactis tRNA fractions (containing arginine tryptophan, and valine acceptor activities) and S. cerevisiae tRNA fractions (containing alanine, asparagine, aspartic acid, glutamic acid, glycine, histidine, tryptophan, and valine acceptor activities) were inactive in the tobacco bioassay. Cytokinins have been found only in those tRNA species corresponding to codons beginning with U.

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A single, modified nucleoside in Escherichia coli tRNA, 6-(3-methyl-2-butenylamino)-2-methylthio-9-#-D-ribofuranosylpurine, has been desulfurized with Raney nickel to afford its probable biosynthetic precursor. The limitations of the reaction at the nucleoside and tRNA levels and its lack of inhibition of the amino acid acceptor activity of tRNA are described.Certain modified nucleosides found in tRNA are also potent plant growth hormones (1). Where these hormones, or cytokinins, occur in tRNA, they occupy only that position adjacent to the 3'-end of the anticodon triplet, and only in tRNA species that respond to codons beginning with uridine (1, 2). Four cytokinins (1-4) have been isolated as components of tRNA (1-3).To facilitate the study of the metabolic function and biosynthesis of one of these modified nucleosides, 6-(3-methyl-2-butenylamino)-2-methylthio-9-,3-D-ribofuranosylpurine (3), a procedure has been developed for hydrogenolysis of the methylthio groups of 3, where it occurs in Escherichia coli tRNA.The identification of cytokinins as components of tRNA has prompted considerable research into the metabolic importance of these compounds. Studies on E. coli tRNA, for example, indicated that cytokinins were located adjacent to the 3'-end of the anticodons in tyrosine (4) and phenylalanine (5) tRNAs, and were also present, presumably adjacent to the anticodon, in cysteine, tryptophan, serine, and leucine tRNAs (6, 7). Gefter and Russell (8) also presented evidence that the cytokinins were important in the specific binding of tRNA to the appropriate ribosome-bound codon triplets, and Fuller and Hodgson (9) hypothesized the importance of this group to proper codon-anticodon interaction in protein synthesis.Peterkovsky (10) and Hall (11) showed that the mevalonate-requiring bacteria Lactobacillus acidophilus and Lactobacillus plantarum rapidly incorporated [14C Imevalonate into the side chain of 6-(3-methyl-2-butenylamino)-9-j3-Dribofuranosylpurine (4) in tRNA. Hall (12-14) later showed that tRNA, which had been partially modified by treatment with permanganate, incorporated [14C]A2-isopentyl pyrophosphate to a greater extent than unmodified tRNA. In addition, Miura and Miller (15) demonstrated that the fungus Rhizopogon roseolus contained the enzymes necessary to convert exogenous 6-(3-methyl-2-butenylamino)purine (4, purine portion) to the hydroxylated derivative 2. Prior to the identification of 3 as a component of E. coli tRNA, Goodman et al. (4) showed that the nucleoside adjacent to the anticodon of E. coli tRNATVT, now assumed to be 3, could be labeled with both [n5S]sulfate and [methyl-14C1-methionine. Rosenberg and Gefter (16) presented evidence consistent with an iron dependence for the in vivo formation of 3 from 4 in tRNA. After the identification of 3 as a tRNA component (17)(18)(19), Harada et al. (19) suggested that this nucleoside must be formed by the modification of 4 in one or two steps.To test for the presence of an enzyme(s) that would modify 4 to 3 at the tRNA level, and to...

Chromatographic systems that employ an active combination of molecular sieve and adsorption chromatography are useful for the rapid purification of enzymes. Such systems, called sievorptive chromatography, will separate either strongly interacting macromolecules or similar macromolecules under conditions of maximal resolution. If ap-