Spectrophotometrical and immunochemical studies on the conformational changes in poly(dG-dC).poly(dG-dC) after modification by 4-hydroxyaminoquinoline 1-oxide

Poly(dG-dC).poly(dG-dC) was modified by the reaction with 4-hydroxyaminoquinoline 1-oxide (4HAQO) in the presence of seryl-AMP. The conformations of 4HAQO-modified poly(dG-dC).poly(dG-dC) and of poly(dG-dC).poly(dG-dC) were studied by circular dichroism spectra under various salt concentration conditions. 4HAQO residues to guanine bases are inefficient in inducing the transition of poly(dG-dC).poly(dG-dC) from B-form to Z-form conformation. We have elicited monoclonal antibodies against 4HAQO-poly(dG-dC).poly(dG-dC). They were characterized using enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA) and binding to supercoiled DNA. These antibodies reacted with 4HAQO-poly(dG-dC).poly(dG-dC) specifically but not with 4HAQO-modified DNA or poly(dG).poly(dC). However, they cross-reacted with N-acetoxy-2-acetylaminofluorene-modified poly(dG-dC).poly(dG-dC) in Z-form conformation. These monoclonal antibodies may recognize a unique conformation in poly(dG-dC).poly(dG-dC) after 4HAQO modification.     

Real-Time Monitoring of Slow-Wave Sleep by Electroencephalogram Variance

Based on statistical variance as an index of electroencephalogram (EEG) parameters, we monitored slow-wave sleep in both humans and rats in real time and on-line with a widely used personal computer. This EEG variance method may be a useful tool to carry out biological rhythm research, including sleep studies.     

Prevalence of spotted fever group rickettsia antibody in Apodemus speciosus captured in an endemic focus in Miyazaki Prefecture, Japan

Fourteen of Apodemus speciosus (large Japanese field mouse) were captured near the place where one of the patients with spotted fever group rickettsiosis had been infected, in Takaoka town, Miyazaki Prefecture. In the town, four human cases were reported. All of the mice had antibodies against Rickettsia japonica and R. montana. The incidence of the antibody was significantly higher in Apodemus mice in the area than in those from nonendemic area.     

Evaluation of the timing to reduce the initial dose of antithyroid drugs in patients with Graves' disease.

The present study was undertaken to evaluate whether the normalization of the serum TSH level in a supersensitive assay during the initial treatment with antithyroid drugs (ATD) is a useful indicator for the reduction of the initial dose of ATD in 50 patients with hyperthyroidism due to Graves' disease. The initial dose of ATD was continued until the achievement of the euthyroid state, and was then reduced either before the serum TSH level was in the normal range in 9 of 29 patients treated with methimazole (MMI) (group MMI-1) and 8 of 21 treated with propylthiouracil (PTU) (PTU-1), or after the serum TSH level was in/above the normal range in 20 of 29 treated with MMI (MMI-2) and 13 of 21 treated with PTU (PTU-2). Although there were no significant differences in age, sex, thyroid function, prevalence of autoantibodies, goiter size, duration of the disease or the initial and modified doses of ATD, the mean durations of the administration of the initial dose of ATD in MMI-2 and PTU-2 were significantly longer than those in MMI-1 and PTU-1, respectively. As a result, 4 (44%) in group MMI-1, 20 (100%) in MMI-2, 2 (25%) in PTU-1 and 7 (54%) in PTU-2 developed low free T4 levels, and 1 (11%) in MMI-1, 15 (75%) in MMI-2 and 3 (23%) in PTU-2 developed low free T3 levels. Serum TSH levels increased over the normal range in 3 (33%) in MMI-1, 18 (90%) in MMI-2 and 5 (39%) in PTU-2.(ABSTRACT TRUNCATED AT 250 WORDS)     

IgE-mediated histamine release in rat basophilic leukemia cells: receptor activation, phospholipid methylation, Ca2+ flux, and release of arachidonic acid

Stimulation of IgE receptors on rat basophilic leukemia cells causes a transient rise and fall of methylated phopholipids, Ca²⁺ influx, and release of arachidonic acid previously incorporated into phosphatidylcholine and liberation of histamine. Inhibition of phospholipid methylation by methyltransferase inhibitors, 3-deazaadenosine and homocysteine thiolactone, almost completely blocks the influx of Ca²⁺, and release of arachidonic acid and histamine. Stimulation of immunoglobulin E receptors by antigen releases only [¹⁴C]arachidonic acid but not [¹⁴C]linoleic acid, [¹⁴C]oleic acid and [¹⁴C]stearic acid, all of which were previously incorporated into phospholipids. [¹⁴C]Arachidonate was found to be incorporated mainly into phosphatidylcholine. The phosphatidycholine rich in arachidonate appeared to be synthesized to a considerable extent by the transmethylation pathway. These findings suggest that in rat basophilic leukemia cells, immunoglobulin E receptors, phospholipid methyltransferases, Ca²⁺ ion channel, and phospholipase(s) that cause release of arachidonic acid and the discharge of histamine are associated.     

Phospholipase activation in the IgE-mediated and Ca2+ ionophore A23187-induced release of histamine from rat basophilic leukemia cells

The importance of phospholipase(s) activation in the IgE-mediated and ionophoreinduced histamine release from the rat basophilic leukemia cell line has been examined. The activation of phospholipase(s) as measured by [¹⁴C]arachidonic acid release and the release of histamine both required Ca²⁺ and were temporally parallel. Inhibition of phospholipase(s) activity by the inhibitors mepacrine and α-parabromoacetophenone also correlated with the inhibition of histamine release. [¹⁴C]Arachidonic acid released by the phospholipase(s) was mainly metabolized to prostaglandin D₂. The inhibition of the cyclooxygenase pathway by indomethacin did not affect histamine release. 5,8,11,14-Eicosatetraynoic acid inhibited both histamine and [¹⁴C]arachidonic acid release suggesting an effect not only on the cyclooxygenase and lipoxygenase pathways but also on the phospholipase(s). These results suggest that activation of phospholipase appears to be necessary for histamine release in the rat bosophilic leukemia cells.     

Circular dichroism and the conformational properties of soybean beta-amylase

The conformational properties of soybean β-amylase were investigated by the circular dichroism probe and measurement of enzyme activity. The enzyme exhibited a positive circular dichroism band at 192 nm, a negative band at 222 nm, and a shoulder near 210 nm. Analysis of the spectrum in the far ultraviolet zone indicated the presence of approximately 30% of α helix and 5–10% of β-pleated sheet, the rest of the polypeptide main chain possessing aperiodic structure. In the near ultraviolet reagion, the enzyme protein showed at least six positive peaks at 259, 265, 273, 281, 292, and 297 nm. The positive bands at 292 and 297 nm remained unaltered on acetylation of the enzyme by N-acetylimidazole and were assigned to tryptophanyl chromophores. These bands were affected in intensity in the presence of maltose or cycloheptaamylose, which indicates that some tryptophan residues are situated at the binding sites. The native conformation of soybean β-amylase was found to be sensitive to pH variation (below pH 5 and above pH 10), sodium dodecyl sulfate, guanidine hydrochloride, and heating to 50–55 °C. Complete disorganization of the secondary structure was attained by 6 m guanidine hydrochloride. Sodium dodecyl sulfate was effective in disturbing the tertiary structure of the enzyme but did not affect significantly the secondary structure. Enzymatic inactivation was paralleled by the decrease of circular dichroism bands in the near ultraviolet region as produced by the denaturants. It is concluded that the uniquely folded structure of the enzyme contains some less rigid domains and a rigid core stabilized by hydrophobic interactions, electrostatic interactions, and hydrogen bonds.     

Acceleration of the Decay of Membrane Potential after Energization of Chloroplasts in Intact Zea mays Leaves

The relationship between dissipation of the flash-induced membranepotential across the thylakoid membrane and the high energystate was studied in Zea mays leaves. The dark decay of theflash-induced 515-nm absorbance change was accelerated by shortpreillumination of the leaf. No acceleration of the decay bypreillumination was observed when leaves were incubated in argonor CO² gas or treated with DCMU. These effects of preilluminationand incubation were reversible. The delayed fluorescence from chlorophyll a was reversibly decreasedby incubating leaves in argon or CO² gas, though the modes ofdepression were somewhat different from each other. In leavesincubated in argon or CO² gas, the phase of slow decrease ofthe intensity of prompt fluorescence during illumination reversiblydisappeared. The results suggested that the dissipation of membrane potentialgenerated by a flash was accelerated after the energizationof chloroplasts in leaves, probably by increased H^– permeabilityof the thylakoid membrane. O2 was important in maintaining (indarkness) and forming (under illumination) the high energy statein chloroplasts in intact leaves. (Received October 1, 1980; Accepted December 15, 1980)     

Co-operative response of chemically excitable membrane III. Three-state model

The general three-state model is formulated first, which is the direct extension of the unified two-state model previously formulated (Kijima & Kijima, 1978). In this model, each protomer in a symmetrically interacting system (oligomers or lattices) can take three states, S, R and Q, where S and R states are the same as in the two-state model and Q state is another state either corresponding to a different open-state of ionophore from R open-state or corresponding to another closed state of ionophore. The model has no restriction on the value of Hill coefficient at the midpoint of the dose-response curves in contrast to two-state models. It is applied on GABA sensitive inhibitory synapse of crayfish muscle to account for anomalous behaviour of the membrane in I^? solution.The simplified versions of the above general three-state model are also formulated (simplified three-state model), in which it is assumed that R and Q state are equivalent in regard to the nearest neighbor interaction. By this assumption, R and Q state are collectively treated as state A and mathematical formula obtained on Ising model are applicable on this model. This model is applied on the insect sugar receptor which was shown to be incompatible with the two-state models (Kijima & Kijima, 1980). Further simplification of the above simplified model results in two convenient models: three-state KNF model and three-state MWC model, which have minimum parameters but sufficient to account for most experiments. They give plausible physico-chemical base on the “classical model” in which the existence of both inactive and active ligand-receptor complex is assumed.     

Anti-platelet aggregating and disaggregating activities of 6,9-methano PGI2

Anti-platelet aggregating and disaggregating activities of the chemically stable 6,9-methano prostaglandin I₂ (6,9-methano PGI₂) were investigated. 6,9-Methano PGI₂ inhibited ADP-induced platelet aggregation in PRP from humans, rabbits and rats. 6,9-Methano PGI₂ also inhibited rabbit platelet aggregation induced by ADP, collagen, thrombin, arachidonic acid and 11,9-epoxy-methano PGH₂. Antiaggregating activities of 6,9-methano PGI₂ were 0.3 to 2.0 times greater than those of PGE₁. 6,9-Methano PGI₂ facilitated platelet disaggregation in a dose related manner. Antiaggregating and disaggregating activities of 6,9-methano PGI₂ were markedly enhanced by incubation with the phosphodiesterase inhibitor, theophylline.