Reactivation in vitro of inactive ribosomes from stationary phase Escherichia coli

Scheps, Ruth; Wax, Richard G.; Revel, Michel

doi:10.1016/0005-2787(71)90498-9

Cited by 20 publications

(5 citation statements)

References 10 publications

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“…Under such ribosome-limiting assay conditions, the kinetics of ['4C]valine incorporations by midlogarithmic and stationary phase ribosomes were similar (Fig.4C). As has been noted earlier [4,21], postribosoma1 supernatants from midlogarithmic and stationary phase cells have similar activity (Fig. 4D).…”

Section: Low Activity Of Unfractionated Extracts From Stationary Phassupporting

confidence: 88%

Metabolic Stability of the Two Forms of Initiation Factor IF‐3 in Escherichia coli during the Growth Cycle

Minks

Suryanarayana

Subramanian

1978

European Journal of Biochemistry

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Possible alteration in the ratio of the long and short forms of initiation factor IF-3 (FEBS Lett. 79. 264-275, 1977) during the growth cycle of Escherichia coli was examined. The ratio was found to remain unchanged between the exponential and stationary growth phases. Contrary to an earlier report (Eur. J . Biochem. 29,319 -325,1972), the total amount of IF-3 relative to the ribosome content in stationary phase cells was essentially the same as in midlogarithmic phase cells. The activity of IF-3, assayed after its separation from other initiation factors by chromatography, was also the same in extracts from midlogarithmic and stationary phase cells. The data show that in Escherichia coli the ratio of IF-3/ribosome is maintained constant.The ribosomes themselves have been shown to retain virtually full activity in uitro during this transition indicating that growth-cycle-dependent biochemical modifications of the ribosome do not affect its protein synthetic capacity per se.Ribosomes in Escherichiu coli have recently been shown to undergo certain biochemical modifications during the bacterium's transition from exponential growth into the stationary phase [l-31. These changes include an increase in the acetylation level of protein L12 in 50-S subunits and a rise in the stoichiometry of a high-molecular-weight protein (denoted the A-protein) associated with 3 0 4 subunits [3]. Another growth-related modification reported in the literature involves initiation factor compared the activities of individual initiation factors in extracts from exponential and stationary phase E. coli and found a striking reduction in the level of IF-3 in the stationary phase.Recently we have shown that different laboratories studying IF-3 have been working with two different forms of this protein and that both forms are present in extracts of E. coli [5]. The amino acid sequences of the two forms have been determined [6] and it is clear that they have the same primary structure except for a hexapeptide segment found at the N-terminus of the longer form. The long and short forms have been designated , respectively [5,6].In view of the reported growth cycle dependence of I F-3 we analysed by two-dimensional gel electrophoresis initiation factor preparations from E. coli harvested at different points in the growth cycle to see whether the ratio of IF-3JIF-3, is altered in uiuo. Such an alteration was not found. But to our surprise we noted that the amount of IF-3,+IF-3, in a given amount of ribosomes (as visually estimated from the gels) did not show a decrease in the stationary phase. Since a protein can conceivably be modified in such a manner that its activity is abolished without alteration in electrophoretic mobility, we determined the IF-3 activity in midlogarithmic and stationary phase E. coli cells. Results presented below show that contrary to the earlier report [4], the IF-3 level is stable during the growth cycle of E. coli. MATERIALS AND METHODS Bacteria and Growth ConditionsIn this study E. coli A1 9 and MRE600 (RNAase-Idefi...

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Section: Low Activity Of Unfractionated Extracts From Stationary Phassupporting

confidence: 88%

Metabolic Stability of the Two Forms of Initiation Factor IF‐3 in Escherichia coli during the Growth Cycle

Minks

Suryanarayana

Subramanian

1978

European Journal of Biochemistry

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“…The present work gives support to this hypothesis by showing that the EF-T, and lFzcatalysed splitting of GTP by ribosomes is strongly inhibited by the nucleoside tetraphosphate, in vitro, North-Holland Publishing Company -Amsterdam was measured by the method of Scheps et al [21].…”

Section: Introductionsupporting

confidence: 74%

Inhibition of in vitro protein synthesis by ppGpp

1972

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“…The results of our examination of the inability of stationary phase extracts to stimulate polyphenylalanine synthesis is not unlike that reported for heterotrophic systems (9,10,15,23,24). It was found that a shift in ribosomal distribution occurred from polysomes to primarily inactive 70S monomers and an increase in 50 and 30S ribosomal subunits (9,21,22) and also possibly a reduction in initiation factors in stationary phase extracts (25).…”

Section: Discussioncontrasting

confidence: 56%

Heterotrophic Nature of the Cell-Free Protein-Synthesizing System from the Strict Chemolithotroph, Thiobacillus thiooxidans

Amemiya

Umbreit

1974

J Bacteriol

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A cell-free protein-synthesizing system prepared from the strict chemolithotroph, Thiobacillus thiooxidans, was similar to that of heterotrophs. The poly-U directed system had a temperature optimum of 37 C, but in the presence of spermidine (3 mM) the optimum shifted to 45 C. Although growth of the chemolithotroph occurs only in acid conditions, the pH optimum for the cell-free system was pH 7.2. The endogenous-directed activity in the presence or absence of spermidine was maximal at pH 7.8. Spermidine had a stimulatory effect; however, this effect was dependent on the magnesium and tris(hydroxymethyl)aminomethane (Tris) concentrations. At low Tris concentrations (10 mM), spermidine (3 to 5 mM) could completely replace magnesium. When the Tris concentration was increased (50 mM), spermidine could not replace magnesium. Supernatant and ribosomal fractions from T. thiooxidans were exchanged with those of Bacillus thuringiensis and Escherichia coli, and the ribosomal fraction from the chemolithotroph gave good to moderate stimulation when exchanged with the supernatant from the heterotrophs. On the other hand, the supernatant from T. thiooxidans gave good stimulation when mixed with ribosomes from B. thuringiensis but poor activity with ribosomes from E. coli. Both supernatant and ribosomal fractions prepared from stationary phase extracts of T. thiooxidans were inactive in the cell-free system.

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Reactivation in vitro of inactive ribosomes from stationary phase Escherichia coli

Cited by 20 publications

References 10 publications

Metabolic Stability of the Two Forms of Initiation Factor IF‐3 in Escherichia coli during the Growth Cycle

Metabolic Stability of the Two Forms of Initiation Factor IF‐3 in Escherichia coli during the Growth Cycle

Inhibition of in vitro protein synthesis by ppGpp

Heterotrophic Nature of the Cell-Free Protein-Synthesizing System from the Strict Chemolithotroph, Thiobacillus thiooxidans

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