Conjugal Transfer of Plasmid-Borne Multiple Antibiotic Resistance in Streptococcus faecalis var. zymogenes

A strain of Streptococcus faecalis var. zymogenes, designated JH1, had high-level resistance to the antibiotics streptomycin, kanamycin, neomycin, erythromycin, and tetracycline. These resistances were lost en bloc from approximately 0.1% of cells grown in nutrient broth at 45 C. The frequency of resistance loss was not increased by growth in the presence of the "curing" agents acriflavine or acridine orange, but after prolonged storage in nutrient agar 17% of cells became antibiotic sensitive. Covalently closed circular deoxyribonucleic acid (DNA) molecules were isolated from the parental strain and from antibiotic-sensitive segregants by using cesium chloride-ethidium bromide gradients. DNA molecular species were identified by using neutral sucrose gradients. Strain JH1 contained two covalently closed circular DNA species of molecular weights 50 x 10(6) and 38 x 10(6). An antibiotic-sensitive segregant, strain JH1-9, had lost the larger molecular species. A second sensitive segregant, strain JH1-5, had also lost the larger molecular species but a new molecular species of approximate molecular weight 6 x 10(6) was present. The antibiotic resistances that were curable from the parental strain were transferred to antibiotic-sensitive strains of S. faecalis and to strain JH1-9, during mixed incubation in nutrient broth at 37 C. Data to be described are interpreted to suggest that the transfer is by a conjugal mechanism. Analysis of the plasmid species in recipient clones showed that all had received the plasmid of molecular weight 50 x 10(6). Strain JH1-5 was not a good recipient. Analysis of one successful recipient clone of JH1-5 revealed that it had gained the 50 x 10(6) molecular weight plasmid but lost the 6 x 10(6) molecular weight species. These data are interpreted to mean that the multiple antibiotic resistance is borne by a transferable plasmid of 50 x 10(6) molecular weight, and that in clone JH1-5 this plasmid suffered a large deletion leaving only a 6 x 10(6) remnant which was incompatible with the complete replicon.     

Studies on the fractionation of total transfer ribonucleic acid and purification of an alanine transfer ribonucleic acid from chick embryo

Chick embryo tRNA, prepared by a simple large-scale method, was fractionated on three different ion-exchange columns. In all cases simple chromatographic patterns for various tRNA species were observed, indicating the presence of only a few major species of tRNA for each amino acid. By repeated chromatography one species of alanine tRNA was purified to approx. 80% purity. T₁ ribonuclease digest of this purified tRNA gave a simple chromatographic pattern. Because of the simplicity of the method of preparation of tRNA from this readily available source and the presence of only a few species of tRNA for each amino acid, chick embryo is suited for the study of tRNA and its various functions in higher systems.     

The effect of adrenocorticotrophin on protein degradation in rat adrenal gland and liver

The rate of adrenal protein degradation appears to be slower in rats to which ACTH (adrenocorticotrophin) has been chronically administered. As measured by the exponential decay of radioactively labelled adrenal protein in vivo, the mean half-lives of total protein and of mitochondrial, microsomal and 18000g-supernatant protein were significantly longer in ACTH-treated animals. Experiments in which either [(3)H]leucine or NaH(14)CO(3) was used to label proteins showed that of the fractions studied, the effect on mitochondrial protein degradation was most pronounced. The half-lives of the same subcellular fractions in rat liver were not affected by ACTH. The possibility that the results might have been caused by changes in radioisotope reutilization and pool size is discussed.     

Studies on the macronucleus of Paramecium aurelia

Following specific incorporation of tritiated thymidine into the DNA of the macronucleus of Paramecium aurelia for varying periods of time the vast majority of silver grains in electron microscope autoradiographs are associated with the area containing the small bodies and matrix. At best only 5% of the silver grains overlying the macronucleus are attributable to the large bodies. This is consistent with the view that the large bodies are the nucleoli of the macronucleus.     

Spectral Studies of Iron Coordination in Hemeprotein Complexes: Difference Spectroscopy below 250 mμ

In order to evaluate the feasibility of observing the spectral behavior of protein groups in the coordination sphere of the iron in hemeproteins, criteria are developed to determine whether or not the application of difference absorption spectroscopy to the study of complex formation will be successful. Absolute absorption spectra, 300-1100 mmu, from bacterial catalase complexes are displayed, and the infrared bands correlated with magnetic susceptibility values of similar complexes of other hemeproteins. Dissociation constants for the formation of cyanide and azide complexes of metmyoglobin, methemoglobin, bacterial catalase, and horseradish peroxidase are given. Difference spectra, 210-280 mmu, are displayed for cyanide and azide complexes of these hemeproteins. A band at 235-241 mmu is found in the difference spectra of all low-spin vs. high-spin complexes. The factors which favor the assignment of this band to a transition involving a histidine residue are presented.