Binding of Putrescine and Spermidine to Ribosomes from Escherichia coli

Abstract

1. The binding of putrescine and spermidine to free 70-S ribosomes has been studied by equilibrium dialysis a t 4" C in the presence of physiological concentrations of K+ and Mg2+. The number of binding sites for each polyamine is approximately 800, and the association constants for putrescine and spermidine are 80 and 500 mM-l, respectively. I n the case of spermidine there is evidence that a small fraction of the sites (3-50/, of the total) have an affinity for spermidine that is about 10 times higher.2. Competition experiments suggest that the two polyamines bind a t closely related sites on the ribosome.3. The binding sites for spermidine are distributed in proportion between the two ribosomal subunits.4. The 820, of free 70-S ribosomes (62 8 ) is unaffected in the presence of physiological concentrations of the two polyamines. 5. The effect of pressure in promoting dissociation of 70-S ribosomes to subunits is reduced by putrescine and spermidine.Polyamines are found in a wide variety of organisms and they have been implicated in a considerable number of biochemical reactions. At physiological pH values their primary and, when present, secondary amino groups are protonated and particular attention has been paid to the possibility of ionic interaction with the negatively charged phosphate groups of nuclcic acids (reviewed in [1,2]). However, understanding of their precise role is still meagre.Escherichia coli contains putrescine (1,4-diaminobutane) and spermidine (1-aminopropyl-1,4-diaminobutane). Small amounts of the monoacetylated derivatives of both polyamines are also present [3]. Their pathways of biosynthesis have been elucidated [I], and E. coli also possesses active uptake systems for both molecules [4]. I n 1960 Cohen and Lichtenstein [5] showed that a large fraction of the polyamines of E . coli could be isolated in association with the ribosomes, and that spermidine in particular could substitute for Mg2+ in preserving the structure of the 70-5 particle. However, it subsequently became clear that the concentrations of mono-and divalent cations in the buffer used for the fractionation of extracts determine the proportions of the polyamines that remain associated with the riboso-31.

Stabilization of Ribosomal RNA of the Small Subunit by Spermidine in Staphylococcus aureus

“…We have shown that the presence of spermidine is crucial for maintaining the correct folding of the helix (h44) of 16S rRNA near the decoding center ( Figure 2 ). Spermidine is a polyamine known to stabilize the folding of RNA molecules including rRNA ( Cohen and Lichtenstein, 1960 ; Weiss and Morris, 1970 ; Cohen, 1971 ; Hardy and Turnock, 1971 ; Turnock and Birch, 1973 ). The upper part of h44 was found as one of the preferred sites for the binding of polyamines ( Amarantos et al, 2002 ) and was particularly observed in the 70S crystal structure from E. coli ( Noeske et al, 2015 ).…”

“…Recent study has shown the importance of magnesium ions on structure stability of the 30 S from E. coli ( Jahagirdar et al, 2020 ). Moreover, numerous studies have led to the conclusion that polyamines, which are present in all types of cells, can stabilize the structure of the ribosome ( Zillig et al, 1959 ; Cohen and Lichtenstein, 1960 ; Stevens, 1969 ; Weiss and Morris, 1970 ; Cohen, 1971 ; Hardy and Turnock, 1971 ; Turnock and Birch, 1973 ). It was shown that E. coli cells grown in the absence of polyamines contained a large portion of defective 30 S particles ( Echandi and Algranati, 1975 ).…”

Subunit association defects in Escherichia coli ribosome mutants lacking proteins S20 and L11

“…The resulting wild-type subunits were found to have a normal complement of ribosomal proteins and rRNAs (see below) but proved to require higher concentrations of Mg2+ ions in order to reassociate and to give rise to ribosomal monomers particularly sensitive to the hydrostatic pressure induced by centrifugation (see Results). Since polyamines are known to counteract the hydrostatic-pressure-induced dissociation of ribosomes [34], our finding can be explained by the efficient removal of ribosome-bound polyamines caused by the dialysis and the centrifugation in high-salt buffer [35].…”

The role of 16S rRNA in ribosomal binding of IF-3

“…The ribosomal binding of erythromycin and the peptidyltransferase activity of the 50S subunit are also inhibited (Teraoka and Tanaka, 1973). The 70S ribosomes have been estimated to contain about 800 binding sites for each polyamine, although the binding affinities for the three polyamines are greatly different (Stevens, 1969;Turnock and Birch, 1973). In addition, it is known that the polyamines interact with the rRNA moiety of ribosomes (Turnock and Birch, 1973).…”