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Biomedical subjects

C E Freda

Publications and source records attributed to C E Freda.

8 recordsLinked to original sources
PubMed1975

The binding of polyamines and of ethidium bromide to tRNA.

The binding of spermidine and ethidium bromide to mixed tRNA and phenylalanine tRNA has been studied under equilibrium conditions. The numbers and classes of binding sites obtained have been compared to those found in complexes isolated by gel filtration a low ionic strength. The latter complexes contain 10-11 moles of either spermidine or ethidium per mole of tRNA; either cation is completely displaceable by the other. In ethidium complexes, the first 2-3 moles are bound in fluorescent binding sites; the remaining 7-8 molecules bind in non-fluorescent form. At least one of the binding sites for spermidine appears similar to a binding site for fluorescent ethidium. Similar results are found with E. coli formylmethionine tRNA. Spermine, in excess of 18-20 moles per mole tRNA, causes precipitation of the complex. Putrescine does not form isolable complexes with yeast tRNA and displaces ethidium less readily from preformed ethidium-tRNA complexes. Under equilibrium conditions, in the absence of Mg++, there are 16-17 moles of spermidine bound per mole of tRNA as determined by equilibrium dialysis. Of these, 2-3 bind with a Ksence of 9 mM Mg++, the total number of binding sites is decreased slightly and there appears to be only one class of sites with a Ka = 600 M(-1). Quantitatively similar results are obtained for the binding of spermidine to yeast phenylalanine tRNA. When the interaction between ethidium bromide and mixed tRNA is studied by equilibrium dialysis or spectrophotometric titration, two classes of binding sites are obtained: 2-3 molecules bind with an average Ka = 6.6 x 10(5) M(-1) and 14-15 molecules bind with an average Ka = 4.1 x 10(4) M(-1). Spermidine, spermine, and Mg++ compete effectively for both classes of ethidium sites and have the effect of reducing the apparent binding constants for ethidium. When the binding of ethidium is studied by fluorometry, there are 3-4 highly fluorescent sites per tRNA. These sites are also affected by spermidine, spermine and Mg++. Putrescine has little effect on any of the classes of binding sites. These data are consistent with those found under non-equilibrium conditions. They suggest that polyamines bind to fairly specific regions of tRNA and may be involved in the maintenance of certain structural features of tRNA.

Binding Sites↗
PubMed1971

System of double infection between vaccinia virus and mengovirus.

When L cells are simultaneously infected with vaccinia virus and mengovirus, double interference in the replication of both viruses is observed. Superinfection of vaccinia virus-infected cells by mengovirus during the first 5 hr of infection reduces vaccinia virus yields to between 1 and 3% of controls. The yields of mengovirus are reduced to between 1 and 16% of controls, depending upon the time of superinfection. The replication of vaccinia deoxyribonucleic acid is not inhibited by mengovirus; it is only delayed. On the other hand, vaccinia multiplication severely hinders the replication of mengovirus ribonucleic acid. The double-infected system, at early times, synthesizes proteins that resemble those synthesized in the vaccinia virus-infected cells. Later in infection, however, the pattern is switched to proteins synthesized by mengovirus-infected cells. Possible mechanisms for this double interference in multiplication are discussed.

Animals↗
PubMed1968

T6r+-induced proteins and nucleic acids in Escherichia coli infected in the presence of streptomycin.

Streptomycin does not strongly inhibit T-even phage multiplication in the streptomycin-susceptible polyauxotroph, Escherichia coli strain T(-)H(-)U(-). The relatively slight inhibition, observed earlier, on production of late proteins has now been studied further. The phage-induced ribonucleic acid, synthesized in T6 phage infection in the presence of streptomycin, has been characterized by its base composition, size distribution, and behavior in hybridization tests. Comparison of these properties to those of control samples, taken during either early or late periods of infection, have not shown any significant differences. Phage-induced proteins, synthesized at different times during infection, were studied by disc-gel electrophoresis. Staining and autoradiography of the patterns of pulse-labeled proteins, formed in the absence and presence of the antibiotic showed only slight quantitative changes in the appearance of early proteins. More marked quantitative effects were detected later in infection. Nevertheless, changes in the mobilities of the different proteins were not observed in the streptomycin-treated cultures at any time after infection, suggesting the absence of gross misreading sufficiently great to alter the distinctive electrophoretic patterns of the extracts. Cells infected and incubated in the presence of the antibiotic were found to contain intact virus particles, as shown by electron microscopy. Such infected cells contained extensive deoxyribonucleic acid pools and did not develop the rounded nucleoids with enclosed dense bodies characteristic of the lethal action of the antibiotic. On the other hand, infected bacteria previously exposed to lethal concentrations of streptomycin were unable to synthesize the early enzymes, deoxycytidylate (dCMP) hydroxymethylase and dihydrofolate reductase, or to make phage deoxyribonucleic acid and phage. Such previously killed cells contained the rounded and clotted nucleoids and were unable to unravel this pathological structure after phage infection.

Autoradiography↗
PubMed1966

Streptomycin and infection of Escherichia coli by T6r+ bacteriophage.

Freda, Celia E. (University of Pennsylvania School of Medicine, Philadelphia), and Seymour S. Cohen. Streptomycin and infection of Escherichia coli by T6r(+) bacteriophage. J. Bacteriol. 92:1670-1679. 1966.-The thymineless, histidineless, uracil-less Escherichia coli 15 THU was shown to be sensitive to streptomycin, dying in patterns comparable to that of strain 15 TAU in the presence or absence of the required amino acid histidine. In the absence of histidine, the antibiotic stimulated ribonucleic acid (RNA) synthesis without a detectable inhibition or stimulation of deoxyribonucleic acid (DNA) synthesis. In the presence of streptomycin (40mug/ml) under conditions of multiple infection with T6r(+), lysis of THU occurred 1 hr earlier than did the control, having produced about one-third as much DNA and phage as did the control. In the absence of histidine, thereby preventing synthesis of phage DNA, accumulation of virus-induced RNA was similar for about 30 min in control and streptomycin-treated systems. In the presence of the antibiotic, however, the infected cells accumulated about 50 to 70% more RNA than did the control after 90 min. Nevertheless, the turnover of RNA was not detectably affected by streptomycin. The rate of production and final amount of deoxycytidylate hydroxymethylase, as well as the cut off time of synthesis of this enzyme, were scarcely affected by streptomycin. The beginning of DNA synthesis was delayed about 3 to 4 min by the antibiotic. The incorporation of histidine in infected cells was unaffected for 10 min and was only about 10% less than the control at 70 min. Lysozyme production began at about 10 min in control and antibiotic-treated systems, continued at essentially similarly increasing rates for 20 min, but stopped abruptly in the streptomycin-treated cells despite continuing protein synthesis. With the exception of lysozyme, the production of phage-specific polymers in a streptomycin-sensitive bacterium was only slightly affected by the antibiotic.

Carbon Isotopes↗
PubMed1966

Nature of ribonucleic acid stimulated by streptomycin in the absence of protein synthesis.

Freda, Celia E. (University of Pennsylvania School of Medicine, Philadelphia), and Seymour S. Cohen. Nature of ribonucleic acid stimulated by streptomycin in the absence of protein synthesis. J. Bacteriol. 92:1680-1688. 1966.-The ribonucleic acid (RNA) synthesized in a thymineless, arginineless, uracil-less Escherichia coli strain 15 in the absence of arginine was characterized by sucrose density gradient centrifugation. About 60% of this RNA had sedimentation rates in the range between 4S and 16S, and the remainder was comprised of the 23S and 16S ribosomal components. On addition of streptomycin for 1 hr in the absence of the amino acid, there was an inhibition of synthesis of material of 4S to 16S, whereas 16S RNA was slightly stimulated. Between 1 and 3 hr after addition of the antibiotic, during the precipitous killing of the bacteria in the arginine-deficient culture, the synthesis of 16S ribosomal RNA was specifically and sharply stimulated.

Arginine↗