Polyamines and the Accumulation of Ribonucleic Acid in Some Polyauxotrophic Strains of Escherichia coli

Abstract

The cellular accumulations of polyamines and ribonucleic acid (RNA) were compared in the polyauxotrophic mutants of Escherichia coli strain 15 TAU and E. coli K-12 RCrel met-leu-. Putrescine, spermidine, and their monoacetyl derivatives were the main polyamines in both strains, when grown in glucose-mineral medium. No significant degradation of either "4C-putrescine or '4C-spermidine was found in growing cultures of strain 15 TAU, which requires thymine, arginine, and uracil for growth. Experiments with this organism showed that in a variety of different incubation conditions, which included normal growth, amino acid starvation, inhibition by chloramphenicol or streptomycin, or thymine deprivation, a close correlation was seen between the intracellular accumulation of unconjugated spermidine and RNA. In the presence of arginine, the antibiotics stimulated the production of putrescine and spermidine per unit of bacterial mass. Deprivation of arginine also resulted in an increase in the production of putrescine per unit of bacterial mass, most of which was excreted into the growth medium. However, in this system the antibiotics reduced the synthesis of putrescine. Furthermore, streptomycin caused a rapid loss of cellular putrescine into the medium. The latter effect was not seen in anaerobic conditions or in a streptomycin-resistant mutant of 15 TAU. Methionine added to the growth medium of growing TAU not only markedly increased the total production of spermidine, but also increased both the intracellular concentration of spermidine and the accumulation of RNA. Exogenous spermidine extensively relaxed RNA synthesis in amino acid-starved cultures of 15 TAU. Analysis in sucrose density gradients showed that the RNA accumulated in the presence of spermidine was ribosomal RNA. Cells of E. coli K-12 RCrel met-leu-, grown in a complete medium, had approximately the same ratio of free spermidine to RNA as did strain 15 TAU. However, the relaxed strain showed a much lower ratio of putrescine to spermidine than the stringent 15 TAU. Omission of methionine stopped spermidine synthesis and markedly increased both the intracellular accumulation and the total production of putrescine. It seems that a high intracellular level of spermidine acts as a feedback inhibitor in the biosynthesis of putrescine in this strain. The hypothesis that the intracellular concentration of polyamines may participate in the control of the synthesis of ribosomal RNA in bacteria is discussed. The wide distribution of natural polyamines in Tabor and Tabor (25). Models have been deliving organisms has raised the problem of their veloped (11) to explain the binding of spermidine physiological function. It has long been known and spermine to deoxyribonucleic acid (DNA). that polyamines are essential growth factors for The strong affinity of polyamines to nucleic acids several microorganisms. It has been shown that and their occurrence in ribosomal fractions (3, 19, polyamines are able to stabilize membrane struc-27) suggest that polyamines serve as cati...

In Vivo Suppression of Coding Associated with Bacteriophage-Induced S -Adenosylmethionine Hydrolase

“…Limitation of the supply of SAM by T3-SAMase would presumably limit the de novo synthesis of spermidine. Although the in vivo role for polyamines in ribosome stabilization is not known, Cohen et al (25) suggested that they may be necessary adjuncts to normal ribosome function. The possible effect of decreased spermidine levels has not been investigated in the present studies.…”

Spermidine-Deoxyribonucleic acid interaction in vitro and in Escherichia coli

“…DNA isolation. E. coli strain 15 TAU was grown to late log phase in 100 ml of synthetic medium (28) The cells were washed twice in 70 ml of 20% sucrose-1 mM ethylenediaminetetraacetic acid (EDTA) as before. The spheroplasts were suspended in 1.2 ml of the sucrose-EDTA solution and lysed by gentle mixing with 23 ml of 1 mM EDTA, 0°C, containing 0.25 tg of ribonuclease (RNase) B (Sigma, preheated, 80°C, 15 min) per ml.…”

Affinity-chromatographic purification of S-adenosyl-l-homocysteine hydrolase. Some properties of the enzyme from rat liver

“…It was ninhydrin-positive and migrated as a single band on paper electrophoresis with two buffer systems (0.1 M-citrate/NaOH, pH4.3; 50mMglycine/NaOH, pH 11.0). Staining with ninhydrin (Raina et al, 1967) showed the presence of one reactive amino group per adenosine derivative. The molecular weight determined by mass spectrometry (Jeol JMS D-300) was 339, corresponding to the expected value.…”