Occurrence of 3-acetamido-3,6-dideoxy-4-O-miethyl-L-glucose in the O-polysaccharide of Vibrio anguillarum

The amino sugar 3-acetamido-3,6-dideoxy-4-O-methyl-L-glucose has been characterized in the O-polysaccharide obtained from the bacterial lipopolysaccharide of Vibrio anguillarum. The identity of 1,2,5-tri-O-acetyl-3-acetamido-3,6-dideoxy-4-O-methyl-L-glucitol has been established by electron-impact and chemical-ionization mass spectrometry and by a labeling procedure.

Structural studies of the class II histocompatibility antigens of the ACI rat

The class II antigens of the ACI rat were studied using both conventional alloantisera and monoclonal antibodies. By sequential immunoprecipitation experiments and cell binding studies, alloantisera were shown to contain antibodies to both the RT1.B and the RT1.D gene products. Using one- and two-dimensional gel electrophoresis, the structures of these gene products were shown to be distinguishable. The importance of these differences for the immune response and antigen presentation is discussed.

A 105 000 dalton antigen of transformed mouse cells is a stress protein

Antisera prepared in mice against syngeneic spontaneously transformed AL/N cells (anti-TAL/N serum) identified a number of protein antigens synthesized by simian virus 40 (SV40) transformed cells, among which was a protein with a molecular mass of 105 000 daltons (p105). Of these transformed cell antigens which were immunogenic in a syngeneic system, only p105 was detected in primary mouse kidney cell cultures, p105 isolated from normal and transformed mouse cells was demonstrated to be identical by two-dimensional gel analysis. Relatively small amounts of p105 were synthesized in quiescent primary cultures, while the protein was actively synthesized in SV40-infected as well as in proliferating mouse kidney cells, and its synthesis in quiescent cells could be induced by subjecting the cultures to glucose starvation or heat-shock treatment. Immunofluorescent staining and cellular fractionation showed that p105 is normally localized to cytoplasmic structures. The results suggest that the expression of p105 is intimately associated with the metabolic state of the cell.

Structural investigation of the lipopolysaccharide core isolated from a virulent strain of Vibrio ordalii

The structure of the core oligosaccharide of Vibrio ordalii has been investigated. The studies involved the use of nuclear magnetic resonance, methylation analysis, partial hydrolysis with hydrochloric acid, nitrous acid deamination, partial hydrolysis with sulfuric acid, Smith degradation, and oxidation with chromium trioxide. As a result of these studies the following structure is proposed.[Formula: see text]

Selective inhibitory effects of thyroid hormones on estrogen-induced protein synthesis in chick embryo liver

We have investigated the effect of thyroid hormones on estrogen-induced responses in embryonic chick liver. Administration of thyroid hormones inhibits estrogen induction of vitellogenin, as well as of apoprotein-II of very low density lipoprotein (VLDL apo-II). A proportionate decrease in the concentration of hepatic salt-soluble nuclear estrogen receptor is also observed. In contrast, estrogen stimulation of apoprotein-B (VLDL apo-B) synthesis is relatively resistant to inhibition. The inhibitory effects of the thyroid hormones could be due to increased metabolism and clearance of estradiol-17β in their presence. The relative resistance of estrogen-induced VLDL apo-B synthesis to thyroid hormone inhibition can be explained by its greater sensitivity to low doses of estradiol. In addition, experiments with the antithyroid agent thiourea suggest that, in vivo, estrogen-induced responses could be balanced by the selective inhibitory effects of thyroid hormones.

Isolation and characterization of nuclear envelopes from three variant cell lines of the Shionogi mouse mammary carcinoma: Identification of androgen-dependent peptides

We have isolated and purified, with good yields, nuclear envelopes from an androgen-responsive and from two androgen-unresponsive cell lines of the Shionogi mouse mammary carcinoma after subjecting purified nuclei to DNase at high pH and characterized them morphologically, chemically, and enzymatically. Phase-contrast microscopy revealed the nuclei to be free of cytoplasmic tags and that the nuclear envelopes were isolated as membrane "ghosts." Electron micrographs clearly showed the double-membrane system with nuclear pore complexes which illustrates that the nuclear envelopes were ultra-structurally intact. The nuclear envelopes contained little DNA, low levels of arylesterase or acid phosphatase activity, and undetectable levels of succinate dehydrogenase and 5′-nucleotidase activity. Coomassie blue staining of the nuclear envelope fractions on sodium dodecyl sulfate – polyacrylamide gels for all three cell lines revealed that most of the polypeptides were similar. However, we have identified androgen-dependent peptides of molecular weights 29 000, 32 000, and 34 000 in nuclear envelopes of the androgen-responsive cell line peptide profiles by comparing the nuclear envelopes prepared from the androgen-responsive cell line grown in intact mice, in castrated mice, and in mice which had been injected with testosterone after castration. Further investigation of the androgen regulation of these nuclear envelope peptides may help us understand the molecular mechanisms involved during morphological changes of the nucleus which occur in response to different hormonal environments.

Mechanisms of cholesterol synthesis inhibition by D-glucosamine

The amino sugar D-glucosamine possesses antitumor activity which is thought to depend in part upon its ability to impair cholesterol biosynthesis and damage cellular membranes. The present study examined the effect of glucosamine on acetate utilization for lipid and sterol synthesis in rat C6 glial tumor cells. At cytotoxic concentrations, the amino sugar inhibited [14C]acetate incorporation into nonesterified sterols and lipids but increased the flow of label into cholesteryl esters. A comparison of the rates of acetate utilization for glucosamine metabolism (N-acetylation) and sterol and lipid synthesis suggested that glucosamine might act by competing for a common cytosolic pool of acetyl CoA. The inhibition of lipid and sterol synthesis, however, remained constant over a wide range of extracellular acetate concentrations. These results suggest that, if glucosamine acts by restricting the supply of acetate for these biosynthetic processes, it probably inhibits a step prior to the formation of acetyl CoA. Alternative mechanisms are discussed.

Influence of mercury (II), cadmium (II), methylmercury, and phenylmercury on the kinetic properties of rat liver glutathione peroxidase

The effect of phenylmercury and methylmercury on rat liver glutathione peroxidase (GSH Px) is investigated and compared with that of Hg(II) and with some previously reported results for Cd(II). Analysis of the kinetics of metal binding to the enzyme gives apparent inhibition rate constants: kc = 9.7 mM−1 min−1 for all three mercury compounds and 75 mM−1 min−1 for CdCl2. Glutathione (0.2 mM) protects the enzyme from metal inhibition, decreasing the apparent inhibition rate constants (kc) by 3.6 times for mercury compounds and 4.4 times for CdCl2. K1 for the three mercury compounds is found to be 53 μM. It is unexpected that the same value of K1 exists for all three forms of mercury studied and that inhibition of the enzyme by the metals is a relatively slow process. For Cd(II) the value of K1 is 8.5 μM. It is suggested that inhibition of GSH·Px enzyme activity by cadmium, mercury, and organic mercury salts may not be due to simple complexation of the active site selenium moiety but may be due to a slower process, e.g., an alteration of the enzyme tertiary or quaternary structure.

Endo-type mode of action of (1 → 4)-β-D-glucan cellobiohydrolase from Sclerotium rolfsii

Sclerotium (1 → 4)-β-D-glucan cellobiohydrolase has two types of activities: an endo-type mode of action forming insoluble short fibres from native cotton and its endwise action of removing cellobiosyl units from the nonreducing chain ends of β-1,4-glucans.

Immunological analyses of selected eukaryotic RNA polymerases II

Polyclonal antibodies to native RNA polymerase II of Achlya ambisexualis and Agaricus bisporus were produced in rabbits and in mice. Monoclonal antibodies were produced against the α-amanitin resistant RNA polymerase II of the mushroom A. bisporus. These antibodies were used in comparative cross-reactivity studies with five purified RNA polymerases II (A. bisporus, A. ambisexualis, Saccharomyces cerevisiae, wheat germ, and calf thymus). A method for quantitatively comparing cross-reactivity was developed utilizing an enzyme-linked immunosorbant assay (ELISA). ELIS A comparisons indicated that the two filamentous fungi cross-reacted effectively with one another and depending upon the preparation reacted less effectively with yeast and wheat germ RNA polymerases II. Cross-reactivity measurements were also made by immunoblotting sodium dodecyl sulfate – polyacrylamide separated RNA polymerases II. The mouse anti-A. bisporus RNA polymerase II immunoglobulin G (IgG) and the monoclonal antibody preparations did not react with high molecular subunits of A. bisporus RNA polymerase II. The sera did, however, cross-react with high molecular weight subunits of A. ambisexualis. Similarily, rabbit anti-A. ambisexualis RNA polymerase II IgG reacted only with low molecular weight subunits of A. bisporus RNA polymerase II, but reacted with high molecular weight subunits of A. ambisexualis and wheat germ. Our results indicate differences in the cross-reactivity of native and denatured RNA polymerases II and suggest differences in the tertiary and quaternary organization of the enzymes examined.