Control of glutaminase synthesis in Escherichia coli.
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Biomedical subjects
Publications and source records attributed to F Varricchio.
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Escherichia coli spheroplasts lysed by Brij 58 and deoxycholate were separated into supernatant (S) and membrane fractions by low-speed centrifugation. The membrane fraction was further divided into that which was releasable by deoxyribonuclease (fraction D) and that which was not (M). In the presence of 10(-2)m Mg(2+), the S, D, and M fractions contained, respectively, 60, 20, and 20% of the total cellular ribonucleic acid (RNA). Ribosomal and transfer RNA (rRNA, tRNA) were found in each fraction. The M + D fraction RNA was labeled more by a pulse label. Incorporation of uracil into the D fraction continued only as long as the uptake of exogenous uracil, suggesting that this was a major primary site of RNA synthesis. From pulse-labeled cells, each fraction contained precursor rRNA, and there was a 10S RNA in the M fraction. Ninety per cent of the ribosomal subunits and the ribosomal precursor particles, 26 and 43S, were in the S fraction. Precursor RNA (17S) was found in the 26S precursor particles. The D fraction contained 38% of the polysomes (this does not consider polysomes, if any, of the M fraction) which were labeled four times as much as the supernatant polysomes by a 1-min pulse of uracil. These results are interpreted to mean that new RNA is associated with a cytoplasmic membrane-RNA polymerase-DNA complex.
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The distribution of ribosomes, 30 and 50S subunits and polysomes, at three different growth rates of Escherichia coli strains B and K-12 has been studied. The usual percentage of subunits is about 20%. However, at the lowest growth rate (mu = generations/hour), mu = 0.45 at 30C, the proportion of subunits is about 30%. An exceptional situation exists in K-12 strains growing at maximum growth rate, mu = 1.35, where the percentage of subunits is 45%. Several points of control over ribosome production are thus indicated. It is suggested that "subunit pool" is essentially a reserve. Furthermore, the polysome content when related to deoxyribonucleic acid content varies directly with the growth rate, which indicates the average efficiency of polysomes in protein synthesis does not vary over the range of growth rates tested.
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A group of nitrogen-containing steroids closely related in structure was screened for antibacterial activity, by use of Bacillus subtilis and Sarcina lutea as the test organisms. The most active compounds were cholesterol derivatives containing a tertiary or quaternary nitrogen in, or attached to, the A ring. Similar methyltestosterone or progesterone derivatives were inactive. All of the cholesterol derivatives that inhibited growth were surfactant, and, structurally, they would be classified as cationic detergents. Some of the inactive compounds were surfactant, but, structurally, they would be classified as nonionic detergents. Certain features of the antibacterial activity of one of the active steroids-ND 212 (4-dimethylaminoethyl-4-aza-5-cholesten-3-one methiodide)-were studied. Growth of a culture of B. subtilis containing 5 x 10(7) cells per milliliter was inhibited by 1 mug/ml (1.7 x 10(-6)m) of ND 212. The amount of growth inhibition was directly related to both cell and steroid concentration. Loss of viability was rapid and irreversible. With B. subtilis, cell lysis was observed. With S. lutea grown in C(14)-glucose, ND 212 caused release into the media of up to 25% of the cellular radioactivity. Extensive leakage occurred before loss of viability was observed. At bacteriostatic azasteroid concentrations, there was little leakage. ND 212 was readily bound in large amounts to B. subtilis cells. Inactive azasteroids were bound poorly. C(14)-cholestanone was also bound, whereas C(14)-methyltestosterone and C(14)-progesterone were not bound in significant amounts. At least 50% of the bound C(14)-cholestanone was associated with the membrane fraction.
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To better understand the pathogenesis and slow healing of sickle cell leg ulcers, we analyzed tissues for their content of iron and their immunohistochemical level of basic fibroblast growth factor, transforming growth factor-beta, and fibronectin. Debrided leg ulcer tissue from seven patients with sickle cell anemia were used. All sections stained strongly for basic fibroblast growth factor. The reactions to iron and fibronectin were variable (trace to 4+, 0 to 3+, respectively), and there was weak or negative immunohistochemical staining for transforming growth factor-beta. These findings suggest the possibility that iron and/or a low content of transforming growth factor-beta and fibronectin may play a role in the chronicity of these lesions. Conversely, reducing tissue iron and/or applying transforming growth factor-beta or fibronectin topically may promote the healing of sickle cell leg ulcers.
We have previously reported on the expression of interleukin-4 receptor (IL-4R) on many solid cancer cell lines. In the present study, we have examined the expression of IL-4R on head and neck cancers in situ by immunohistochemistry. Seven primary squamous cell carcinomas of the head and neck region were stained by a monoclonal antibody to human IL-4Rp140 protein (M-57). We report that all squamous cell carcinoma samples were positively stained although at variable intensity with anti-IL-4R antibody. Tumors stained with IgG control did not show any staining. On the other hand, six benign lesions from the same anatomical area showed faint or no staining at all. Three uterine endometrium samples were also negative for the IL-4R expression. In contrast to published contrary results, we did not observe any effect of IL-4 on the proliferation of four squamous cell carcinoma of head and neck (SCCHN) cell lines examined. These results demonstrate that human squamous carcinoma of the head and neck express IL-4R, which could be targeted for diagnosis and therapy by anti-IL-4 receptor antibody fused to toxins or radionuclides or alternatively by IL-4 toxins.