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

Abstract

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...

Stationary phase, amino acid limitation and recovery from stationary phase modulate the stability and translation of chloramphenicol acetyltransferase mRNA and total mRNA in Escherichia coli

“…This trend in translational rates during culture growth is consistent with in vitro translation studies. Extracts obtained from late exponential-phase and stationary-phase E. coli cultures are lower in translational activity when compared to extracts made from early exponential-phase cultures (Minks et al, 1978; Isaksson et al, 1980). Therefore, the decrease in the rate of CAT and total protein synthesis after the exponential growth stage may be due to a decreasing ability of the cell to efficiently translate mRNAs.…”

Functional Subdomains of Yeast Elongation Factor 3

“…Escherichia coli ribosomes were prepared according to the procedures of Minks et al (27). The concentrations of ribosomes and subunits were determined taking 1 A 260 unit to be equivalent to 18.5 pmol of 80 S ribosomes, 24 pmol of 70 S ribosomes, 56 pmol of 40 S subunits, and 27 pmol of 60 S subunits.…”

Function of the repeating homologous sequences in nucleic acid binding domain of ribosomal protein S1

“…Bacterial Strains and Growth Conditions. Escherichia coli strains MRE600 and A19 were grown at 37 °C with vigorous aeration and harvested in midlogarithmic phase as described by Minks et al (1978).…”