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Differentiation of radioactive DNA and RNA deposited on filter paper discs can be accomplished by a relatively simple procedure. RNA can be efficiently removed by incubating the dises, impaled on pins, with 0.2 ml of 0.5 n NaOH for 90 min at 37°C. DNA can be removed after NaOH hydrolysis by treating the discs with 5% TCA for 30 min at 90°C. A correction is necessary to determine the actual amounts of DNA and RNA in order to account for the loss of DNA (13.8%) during the NaOH hydrolysis procedure.

Unfractionated bromegrass mosaic virus RNA and each of its three naturally occurring components, of molecular weight 1.0 × 10 6 , 0.7 × 10 6 , and 0.3 × 10 6 , stimulate the incorporation of amino acids into peptides in an E. coli cell-free incorporating system. Analysis of tryptic digests by paper electrophoresis and chromatography indicated that the products induced by the RNA small component of molecular weight 0.3 × 10 6 , and by the unfractionated RNA were similar.

1. 1. A method is described for the purification of a ribonuclease (polyribonucleotide 2-oligonucleotide transferase (cyclizing), EC 2.7.7.16) from mature, unfertilized eggs of the sea urchin Psammechinus miliaris . The enzyme was purified 19 000-fold with a yield of 48% with the aid of chloroform treatment, (NH 4 ) 2 SO 4 fractionation, and by chromatography on Sephadex G-100, on SE-Sephadex C-50 and on CM-Sephadex C-50. 2. 2. The enzyme had a pH-optimum at 5.3–5.5. 3. 3. The enzyme was heat labile, loosing about 50% of its activity in 3 min at 60° and pH 5.3, 70% of its activity in 3 min at 60° and pH 2.1, and 80% of its activity in 3 min at 40° and pH 9.0. 4. 4. The enzyme hydrolyzed yeast RNA completely to 2′,3′-cyclic nucleotides, and finally hydrolyzed the purine cyclic nucleotides slowly to their corresponding 3′-isomers. 5. 5. The enzyme hydrolyzed guanosine 2′-3′-cyclic phosphate about three times faster than adenosine 2′,3′-cyclic phosphate. 6. 6. The purified enzyme was found to be free of deoxyribonuclease and non-specific phosphodiesterase activity. 7. 7. A molecular weight of about 37 000 was estimated for the enzyme by comparative gel filtration experiments.

From the yeastCandida utilis a compound was isolated which uncouples the process of cell growth and division by accelerating the cell division without influencing outgrowth of cell mass when applied on another population of identical yeasts. This compound accelerated the initiation of DNA synthesis and had no influence on the synthesis of RNA and protein. Moreover, in the presence of division inducing factor the yeasts started multiplication before the content per cell reached the control level. The stimulating effect of division inducing factor was not obvious when the proteosynthesis of yeast cells was inhibited. We concluded that the division inducing factor regulates the formation of a protein which is synthesized in a very small amount and has a role in the initiation of DNA replication.

HeLa cells and L cells were allowed to grow in the presence of l -methioninemethyl- 14 C so that any methylated components of the transfer RNA (tRNA) would be radioactive. The tRNA from these cells was then isolated and a complete analysis was made of the methylated bases present as minor components of the tRNA. For this purpose, the purified RNA was subjected to acid hydrolysis, and the hydrolytic products, including the methylated bases, were partially separated by chromatography on ion-exchange resins. Radioactive (methylated) compounds were then identified by paper Chromatographic analyses and, in some cases, by their chemical characteristics. Methylated bases found in relatively major quantities in both HeLa tRNA and L cell tRNA were: 1-methyladenine, 1-methylguanine, N 2 -methylguan ine, N 2 -dimethylguanine, 7-methylguanine, 5-methylcytosine, and 5-methyluracil (thymine). Methylated bases present in smaller quantities included 1-methylhypoxanthine, 3-methylcytosine, and a base thought to be 3-methyluracil. Evidence was also obtained for the presence in small amounts of 2- O -methylribose residues in both HeLa and L cell tRNA. 5-Hydroxymethyluracil was identified as a minor component of L cell tRNA, but it was not present in HeLa RNA.

Conventional usage of the filter-paper disc technique following in vivo hepatic incorporation of certain labeled pyrimidine and purine RNA precursors resulted in the presence of radioactivity on the filter disc in species other than RNA. Thus the filter paper disc technique, conventionally conducted, may be misleading. To eliminate artifacts, it is proposed that replicate filter discs be processed, one group passing through the usual cold TCA washings only, while a parallel group of replicate discs are heated for 20 min at 90°C in 5% TCA to remove RNA. True RNA radioactivity is the difference of counts from the cold TCA precipitable material and the radioactivity remaining after hot TCA extraction.

The reaction of phosphorothioate with bovine pancreatic ribonuclease at pH 9.0 in the absence of urea led to a derivative, denoted 4PS-ribonuclease, in which two of the four disulphide bonds in the molecule were opened. 4PS-ribonuclease is a unique molecular species, and not a mixture of native and open-chain ribonuclease; this is apparent from electrophoretic studies. By the use of the “diagonal map” technique it has been established unequivocally that the disulphide bonds opened were between half-cystines 3–8 and 4–5, counting from the amino terminus along the ribonuclease chain. 4PS-ribonuclease is fully active enzymically toward RNA, and more active than the native enzyme toward cytidine 2′,3′-cyclic phosphate. It is indistinguishable from native ribonuclease in its immunological reaction with antiserum to ribonuclease, and in its spectrophotometric titration, which shows that three of the six tyrosine phenolic groups in 4PS-ribonuclease titrate abnormally. Unlike native ribonuclease, 4PS-ribonuclease is digested by trypsin, even though at a lower rate than open-chain ribonuclease.

1. RNA-iodo-proteinaceous complexes occur in the tRNA fractions isolated from bovine, ovine, and porcine thyroids. The calf thyroid complex has been partially characterized and contains diiodotyrosine and probably the hormone thyroxine in addition to several other amino acids. 2. Different preparative procedures and techniques capable of disrupting a variety of non-covalent bonds and aminoacyl and peptidyl ester bonds had no obvious effect on the calf thyroid RNA-iodo-proteinaceous complex which remained associated with tRNA. 3. Nucleolytic enzymes can degrade the complex to lower molecular weight products that contain iodine, proteinaceous, and RNA components. However, most of the iodine and proteinaceous material is associated with a relatively high molecular weight, ribonuclease-resistant, RNA-containing core. 4. It is concluded that the tRNA fractions of bovine and probably other mammalian thyroid glands contain a low molecular weight RNA linked to proteinaceous material that contains iodinated amino acids.

The RNA of bromegrass mosaic virus (BMV-RNA) was isolated as three components (L, M, and S, of about 1.0, 0.7 and 0.3·106 mol. wt.) and studied by terminal labeling. Periodate oxidation of the 5′-linked terminus followed by [14C]-dimedone treatment or by reduction with NaB3H4 showed Components L and M to each contain one free 5′-linked end per molecule. Chromatography of ribonuclease T1 hydrolysates of NaB3H4 labeled Components L or M indicated that the two RNA fractions contain the same terminal oligonucleotide; purification and analysis of each oligomer confirmed their identity and permitted tentative identification of the terminal sequence as ... GpCpCpCpA. Component S incorporated less than equimolar quantities of dimedone or NaBH4 after oxidation, and chromatography of 3H terminally labeled material showed some terminal heterogeneity but also much of the same terminal oligonucleotide identified above. Phosphomonoesterase treatment of Component S produced a variety of new periodate-sensitive ends. It was concluded that intact viral RNA (Component L) may be cleaved in a sensitive region (but not at a single internucleotide linkage) to form molecules of Component M which carries the 5′-linked terminus and Component S which usually bears a 3′-phosphate end. Component M may then be further cleaved to form two molecules of Component S, one carrying the original 5′-linked end and one usually terminating in a 3′-phosphate.