Hanna Engelberg

Effect of magnesium and spermine on the aggregation of bacterial and mammalian ribosomes.

Abstract A strikingly different behaviour of Escherichia coli and mouse-liver ribonucleoprotein particles towards Mg 2+ and spermine was observed. Electron micrographs of E. coli ribosomes in 10 mM Mg 2+ showed the characteristic features of a mixed ribosomal population consisting of monomers and small clusters of monomers, “polysomes”. Suspensions of mouse-liver ribosomes in media containing 10 mM Mg 2+ , on the other hand, gave rise to the appearance of extremely large ribosomal aggregates consisting of a minimum of 100 ribosomal particles. These ribosomal aggregates were shown to be held by Mg 2+ alone, the presence of messenger RNA being superfluous. The lowest Mg 2+ concentration at which mouse-liver ribosomes formed aggregates was 5 mM. 30-S and 50-S ribosomal sub-units of E. coli associated into the 70-S form at concentration of spermine of 0.4 mM. Increasing the concentration of spermine to 0.8 mM gave rise to the appearance of 80-S and 100-S ribosomal particles. At 1 mM spermine E. coli ribosomes were reversibly and quantitatively precipitated. Precipitation of mouse-liver ribosomes commenced at 0.1 mM spermine. Quantitative precipitation of mouse-liver ribosomes occurred at a concentration of spermine which was optimal for the preservation of bacterial ribonucleoprotein in the 70-S form (0.4 mM spermine).

Studies on the enzymic oxidation of aminopurines

Abstract 1. 1. 8-Aminopurine is oxidised by mammalian xanthine oxidase along the following pathway: 8-Aminopurine→6-hydroxy-8-aminopurine→2,6-dihydroxy-8-aminopurine It is thus evident that oxidation of all three aminopurines takes a course different from the corresponding hydroxypurines. 2. 2. Adenine and 6-methylaminopurine are oxidised along the same pathway, but all other methyl derivatives of adenine are refractory. 3. 3. 2-Aminopurine is converted by a bacterial enzyme system into 2-amino-8-hydroxypurine. These results are discussed in terms of specific enzyme-substrate complexes.

Studies on streptomycin-dependent bacteria: Uptake of 14C streptomycin by a streptomycin-dependent mutant of Escherichia coli

Uptake of [24C]streptomycin by growing cells and non-proliferating suspensions of a streptomycin-dependent mutant, Sd-4, of Escherichia coli has been studied. The uptake of streptomycin has been shown to be an instantaneous and temperature-independent process. The magnitude of streptomycin uptake has been found to be a function of ionic strength of the medium. Maximal uptake has been observed from salt-free solutions. Increase in ionic strength of the medium brought about a decrease in streptomycin uptake as well as a release of cell-bound streptomycin. Minimum of streptomycin that had to be added to the growth of medium in order to nesure optimal growth of the organism has been found to constitute an approx. 30-fold excess over that actually taken up by the cells. The necessity of adding streptomycin to the growth medium in such a large excess as well as the low streptomycin uptake and release of the cell-bound streptomycin in salt solutions have been explained by a dynamic equilibrium that exists between cell-bound streptomycin and inorganic ions, in all likelihood cations, of the medium i.e. a competition between inorganic ions and streptomycin for common binding sites of the cell. The data indicate that streptomycin is held by the cell of the streptomycin dependent of Escherichia coli through electrostatic interaction of opposite charges, i.e. the positively-charged groups of streptomycin and the negatively-charged groups of cell receptors.

Studies on streptomycin dependent bacteria: Effect of growth in limiting amounts of streptomycin on respiration and fermentation of a streptomycin dependent mutant of Escherichia coli

Respiration and glycolytic of a streptomycin dependent mutant of Escherichia coli Sd-4, grown with optimal and suboptimal amounts of streptomycin were studied. The glycolytic activity of cells grown in limiting amounts of streptomycin was considerably higher than that of cells grown at optimal concentration of streptomycin. On the other hand, the respiration of cells grown with limiting amount of streptomycin was two to five-fold lower than that of cells grown with optimal amounts of sterptomycin. It was found that the deficient cells lacked the carbon monoxide-sensitive pathway of respiration.

Sequence homologies between ribosomal and phage RNAs: A proposed molecular basis for RNA phage parasitism

Abstract An extensive nucleotide sequence homology between the 3′-end of the 16 S ribosomal RNA and segments of bacteriophage MS2 or Qβ RNA is described. In addition, a notable sequence homology of coliphage RNAs with several other segments of ribosomal RNA is shown. The role of bacterial proteins in the recognition of phage RNA, and the resemblance of phage and host RNAs as the molecular basis of RNA phage parasitism is discussed. The problem of quantifying the degree of homology is discussed in the Appendix with a preliminary attempt towards a solution. A relative measure of homology is described, and used to analyze statistically the data obtained.

Inhibition of RNA bacteriophage replication by rifampicin

Abstract Rifampicin has a differential effect on the two steps of MS2 phage replication in vivo : RNA minus strands are synthesized on the parental template at a reduced rate, whereas the synthesis of progeny plus strands is completely inhibited. The effect of the drug is not immediate, rather, a period of approximately 15 minutes is required for the drug to exert a maximal inhibitory effect on both stages of RNA replication. Inhibition of phage RNA replication by rifampicin could be interpreted as a competition of RNA phage replicase and DNA-dependent RNA polymerase for a common protein(s).

Studies on the fate of parental bacteriophage RNA in the host

Abstract MS2 phage particles incapable of forming plaques are capable of RNA injection. The RNA injected by non-viable phage comprises about 90% of the total phage RNA which penetrates the host cells. This RNA is soon after injection converted into all nucleic acid components of the cell. At the same time about 10% of the parental phage RNA was found in the double stranded form, a value which corresponds to the total amount of RNA injected by viable phage, indicating quantitative conversion of this RNA into replicative form, soon after injection.