Isolation, purification and properties of 5 s ribosomal RNA: a new species of cellular RNA

Comb, Donald G.; Zehavi‐Willner, Tova

doi:10.1016/s0022-2836(67)80117-7

Cited by 74 publications

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“…In view of the selective inhibitory action of both drugs on the formation of high-molecular-weight rRNA, it was considered interesting to investigate also the effect of cycloheximide and FU on the formation of low-molecular-weight rRNA. In the present studies, separation of both low-molecular-weight RNA species was obtained by chromatography on Sephadex G-100 (15) and on diethylaminoethyl (DEAE) cellulose at 80 C (2). The results showed that the formation of low-molecularweight rRNA is less inhibited than the formation of its high-molecular-weight counterparts, but considerably more than the formation of tRNA.…”

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confidence: 75%

“…Separation of low-molecular-weight RNA components by chromatography on DEAE cellulose at 80 C. Separation of low-molecular-weight RNA species was also achieved by chromatography on DEAE cellulose at 80 C, as described by Comb and Zehavi-Willner (2). About 500 Mg of low-molecular-weight RNA was applied on a column (10 by 1 cm) and eluted at 80 C, with 150 ml of a linear gradient of 0.25 M to 1.5 M NaCl in 0.02 M tris(hydroxymethyl) aminomethane (Tris)-hydrochloride buffer (pH 7.6).…”

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Effects of Cycloheximide and 5-Fluorouracil on Formation of Low-Molecular-Weight Ribonucleic Acid in Yeast

1969

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The effects of cycloheximide and 5-fluorouracil on the formation of low-molecularweight ribonucleic acid (RNA) in yeast were investigated. Both compounds were found to affect the synthesis of low-molecular-weight RNA of ribosomal origin more than transfer RNA but less than the high-molecular-weight ribosomal RNA. 5-Fluorouracil-containing transfer RNA was separated from normal transfer RNA by chromatography on diethylaminoethyl cellulose at 80 C in the presence of 7 M urea. G-100 (Pharmacia, Uppsala, Sweden) according to the method described by Virmaux et al. (15). About 1,000 to 4,000 jig of RNA dissolved in 2 ml of 0.05 M ammonium acetate buffer (pH 5.0) was applied to a column (180 by 2 cm) of Sephadex G-100 made in the same buffer. Fractions (5 ml) were collected at a flow rate of 35 ml/hr; the first 200 ml of effluent was discarded. The optical density at 260 nm was read, 743 on July 16, 2020 by guest http://jb.asm.org/ Downloaded from on July 16, 2020 by guest http://jb.asm.org/ Downloaded from on July 16, 2020 by guest http://jb.asm.org/ Downloaded from on July 16, 2020 by guest

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Effects of Cycloheximide and 5-Fluorouracil on Formation of Low-Molecular-Weight Ribonucleic Acid in Yeast

1969

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“…The major nucleotide composition of soluble and ribosomal 5 S RNA was identical (Up, 23.4; Gp, 31.6; Cp, 26.8; Ap, 18.2; by per cent). One of the characterizing features of ribosomal 5 S RNA molecule is the absence of any detectable methylated base in it [4,6,7], and its lack of methyl accepting activity as demonstrated in vitro (Tova Zehavi-Willner, unpublished results). Therefore it was interesting to test whether the soluble 5 S RNA also lacked methylated bases.…”

Section: Isolation and Properties Of A Low Molecular Weight Rna (Solumentioning

confidence: 97%

“…It was, therefore, considered to be a universal component of both prokaryotic and eukaryotic ribosomes. While further analysis of the cellular location of 5 S RNA has shown that the molecule attached to each ribosome is always associated with the large ribosomal subunit [6][7][8][9], it was also found on the more mature precursor particles of the 50 S subunit [10][11][12]. No soluble 5 S RNA was detected in the cytoplasm of either prokaryotic or eukaryotic cells, although the existence of minor quantities of such molecules was not excluded [13,14].…”

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The isolation and properties of reticulocyte soluble 5 S RNA

Zehavi‐Willner¹,

Danon²

1972

FEBS Letters

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“…Concentrations were determined optically at room temperatures using an extinction coefficient of 0.80 X 104 per residue at 260 mnu. 11 All optical measurements were made in 0.01 M tris-chloride buffer, pH 7.3, containing 0.3 M NaCl and 0.001 M MgCl2. Optical rotatory dispersion (ORD) and UV absorption measurements employed the Cary 60 spectropolarimeter and Cary 15 spectrophotometer, respectively.…”

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The extent of base pairing in 5S ribosomal RNA.

Cantor¹

1968

Proc. Natl. Acad. Sci. U.S.A.

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Up to the present time most physical studies of the conformation of ribonucleic acids in solution have concentrated on synthetic homopolynucleotides, transfer RNA, or large viral and ribosomal RNA's.1 Synthetic polymers have long been used as models for natural RNA's but the vast majority of compounds currently available have very regular sequences unlike those found in biosynthetic molecules. Thus they do not demonstrate the intrachain folding thought to occur in such molecules as tRNA.2 Studies on natural RNA's have been hindered by their considerable complexity. The paucity of nucleotide sequence information for viral and ribosomal RNA's has made conformation studies on these systems very difficult. The presence of many unusual nucleotides simplified the sequence determination of tRNA's,3-6 but the optical properties of these bases are largely unknown. This complicates attempts to explore the conformation of tRNA's by optical means. What is needed is a simpler system on which various conformational probes can be tested. The discovery of the relatively small 5S ribosomal RNA7-8 and its subsequent sequence determination by Brownlee, Sanger, and Barrell provides such a system.9 This RNA contains 120 nucleotide residues. The only bases which appear are the four normal ones, A, U, C, and G. In this paper we shall describe the results of several preliminary optical studies on 5S ribosomal RNA. These demonstrate that the molecule does indeed contain a large percentage of double-stranded helical regions. At present there is not enough detailed information to permit a likely conformation to be chosen from among the many possibilities which present themselves when the nucleotide chain is folded in three dimensions. But our results suggest an approximate picture of the conformation of 5S RNA which should serve to stimulate further experiments.Materials and Methods.-5S ribosomal RNA was furnished by Drs. K. Hosokawa and M. A. Q. Siddiqui. It was prepared from the total RNA of E. coli strain A-19, deficient in ribonuclease, by a method to be published, using Sephadex column chromatography.10 It was found to be homogeneous by MAK (methylated albumin kieselguhr) column chromatography. Concentrations were determined optically at room temperatures using an extinction coefficient of 0.80 X 104 per residue at 260 mnu. 11 All optical measurements were made in 0.01 M tris-chloride buffer, pH 7.3, containing 0.3 M NaCl and 0.001 M MgCl2. Optical rotatory dispersion (ORD) and UV absorption measurements employed the Cary 60 spectropolarimeter and Cary 15 spectrophotometer, respectively. One-centimeter path length jacketed cylindrical cells were used for all experiments. These were sealed with a serum stopper pierced by a 25-gauge syringe needle to retard evaporation. The temperature inside the sample cell was calibrated using a thermocouple during dummy runs where the cell contained buffer instead of sample. All temperature studies were corrected for the thermal expansion of the solvent.Results.-Three types of optical experiments wer...

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Isolation, purification and properties of 5 s ribosomal RNA: a new species of cellular RNA

Cited by 74 publications

References 19 publications

Effects of Cycloheximide and 5-Fluorouracil on Formation of Low-Molecular-Weight Ribonucleic Acid in Yeast

Effects of Cycloheximide and 5-Fluorouracil on Formation of Low-Molecular-Weight Ribonucleic Acid in Yeast

The isolation and properties of reticulocyte soluble 5 S RNA

The extent of base pairing in 5S ribosomal RNA.

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