Immunochemical subfractionation of a microsomal fraction of rat liver with antibody-coated Staphylococcus aureus cells

The procedure for immunochemical adsorption of vesicles with specific antigen on their outer surfaces was improved. When microsomal vesicles were mixed with Staphylococcus aureus cells coated with the antibody against NADPH-cytochrome c reductase, more than 90% of the enzyme activity was adsorbed on the cell, whereas, only about 10% of the activity was adsorbed on cells coated with the same amount of anti-ovalbumin antibody. NADH-cytochrome c reductase and aldehyde dehydrogenase activities were adsorbed on the cell to the same extent as was NADPH-cytochrome c reductase activity. Under this condition, there was no adsorption of the activities of the marker enzymes of lysosomes and Golgi apparatus, whereas large amounts of the activities of the plasma membrane enzymes were adsorbed. The specific activity of NADPH-cytochrome c reductase in the adsorbed vesicles from the microsomal fractions increased considerably. In contrast, marker enzymes of the Golgi or of the plasma membranes could be enriched in unadsorbed vesicles from the Golgi fractions.     

Metabolism of 3-phosphoglyceroyl phosphate in phosphoenolpyruvate-enriched human erythrocytes

The concentration of 3-phosphoglyceroyl phosphate in erythrocytes was increased by more than 100-fold when red cells were incubated with extracellular phosphoenolpyruvate at 37 degrees C. Since these elevated levels were maintained for 60 min, the metabolism of 3-phosphoglyceroyl phosphate and related compounds could be investigated in phosphoenolpyruvate-treated erythrocytes. 2,3-Bisphosphoglycerate synthesis was not affected by intracellular pH when the 3-phosphoglyceroyl phosphate level was constant but did vary with 3-phosphoglyceroyl phosphate concentration. On the other hand, the relationship between the rate of 2,3-bisphosphoglycerate synthesis and 3-phosphoglyceroyl phosphate concentration was not straightforward. At relatively low concentrations of 3-phosphoglyceroyl phosphate, the observed rate of 2,3-bisphosphoglycerate synthesis agreed with a rate calculated from a formula incorporating kinetic parameters of purified 2,3-bisphosphoglycerate synthase (Rose, Z.B. (1973) Arch. Biochem. Biophys. 158, 903-910). However, at high concentrations of 3-phosphoglyceroyl phosphate, the observed rate of 2,3-bisphosphoglycerate synthesis was lower than the calculated value. The concentration of glucose 1,6-bisphosphate did not increase even when 3-phosphoglyceroyl phosphate was elevated to 200 microM. Elevated levels of intracellular 2,3-bisphosphoglycerate did not inhibit glycolytic activity in these erythrocytes. These results suggest that incubation of erythrocytes with phosphoenolpyruvate is a useful technique to investigate the effect of metabolic perturbations at the intermediate stages of glycolysis.     

Calcium-activated protease in the giant algaChara australis

Summary There is a protease, which is activated by Ca²⁺ (about 100 M), works at neutral pH and exists in the cytoplasm inChara australis. This protease may correspond to calpain, the calcium-activated neutral protease, which has been studied in animal cells. This is the first report showing the existence of a calcium-activated protease in plant cells.     

Synchronous division and the pattern of diel vertical migration of Heterosigma akashiwo (Hada) Hada (Raphidophyceae) in a laboratory culture tank

Heterosigma akashiwo (Hada) Hada (Raphidophyceae) causes red tides in Osaka Bay (Japan). A clonal culture of the alga was grown in a 2 m tall culture tank on a 12: 12 LD cycle to determine patterns of vertical migration and cell division. A specific growth rate of 0.43 ln unit · day⁻¹ was obtained during complete mixing conditions. Under weakly stratified conditions (≈ΔT = 3–4°C/1.5 m), H. akashiwo in the tank grew and showed a similar pattern of vertical migration to that observed in the field for at least 6 days. Cell concentration, mean cell volume, and photosynthetic capacity, estimated by DCMU-induced fluorescence increase of H. akashiwo, were monitored in the stratified tank at 2-h intervals over 24 h at three levels in the water column. Cell ascent began shortly before the light period and vertically swimming cells were smaller in size than those sampled near the bottom of the tank. The cell division cycle and the pattern of vertical migration were phased individually by the light regime and were well synchronized with each other. This synchrony must be due to the interrelation between these two processes or the existence of a clock which controlled endogenous rhythms of both processes and was entrained by a light: dark cycle. The relative increase of fluorescence with DCMU was higher for migrating cells than for non-migrating cells.     

Level of translatable messenger RNA coding for argininosuccinate synthetase in the liver of the patients with quantitative-type citrullinemia

Summary The translation activity of mRNA coding for argininosuccinate synthetase in total RNA extracted from the liver of three patients with quantitative-type citrullinemia was determined using a cell-free translation system. In two patients, the hepatic content of the enzyme was about 20% of the control value, whereas translatable mRNA level for the enzyme was similar to or slightly lower than those of control livers. In the third patient, the enzyme content was about 50% of the control value, and mRNA activity for the enzyme was low normal. These results indicate that at least in the first two patients, the decrease in the enzyme protein is due either to increased degradation of the enzyme or to decreased translation in the patient's liver.     

Triterpenes from Periandra dulcis roots

Three oleanane triterpenes were isolated from the roots of Periandra dulcis,and identified as 3β-hydroxy-25-al-olean-18-en-30-oic acid (periandric acid I), 3β-hydroxy-25-al-olean-12-en-30-oic acid (periandric acid II) and 3-oxo-25-hydroxy-olean-12-en-30-oic acid. The former two compounds (periandric acids I and II) were identical with the aglycones obtained by hydrolysis of periandrin I and II, respectively and the latter one was a new triterpene.     

Concentrations of essential elements after repeated administrations of tin and selenium

To study effects of simultaneous administration of tin (Sn) and selenium (Se) on concentrations of several essential elements, mice were injected with either SnCl2 (ip) or Na2SeO3 (sc), alone or both compounds at a daily dose of 5 mumol/kg each for 12 consecutive days. Mice were sacrificed at 20 h after the last injection and concentrations of Sn, Se, Na, Ca, Zn, P, Fe, K, and Mg in the liver, kidney, spleen, pancreas, testis, seminal vesicle, lung, femoral muscle, and femoral bone were determined. In the control mice, Sn and Se concentrations were the highest in bone (0.69 micrograms Sn and 6.93 micrograms Se/g dry wt). Administered Sn was found to accumulate in all organs except the testis. Among the essential elements determined, Na was the most affected in terms of concentration in the organs and Mg was the least affected element in these organs. Among the organs tested, each elemental concentration in the pancreas was most affected. Simultaneous injections of Sn and Se appeared to keep the correlation coefficients between elements similar to those found in the control mice.     

Regulation by Androgen of Levels of the β Subunit of Nerve Growth Factor and Its mRNA in Selected Regions of the Mouse Brain

Abstract: Our previous studies showed that the concentration of the β subunit of nerve growth factor (β-NGF) in nervous tissues is higher in male than in female mice. To identify the brain regions that are affected by androgens, the amounts of β-NGF protein and its mRNAs were measured in male, female, and castrated male CD-1 mice and testicular feminization mice at 3–4 months of age. Among tissues examined, the hypophysis of males contained the highest average concentration of β-NGF protein. In most regions of the brain, individual levels were more variable in males than in females. However, after the castration, such variations in β-NGF levels disappeared. Average levels of β-NGF protein in males were higher in the cerebellum (eightfold higher), olfactory bulb (12-fold higher), hypothalamus (sixfold higher), and hypophysis (72-fold higher) than thope in corresponding regions of females. No significant differences were observed in levels of β-NGF protein in the hippocampus, cerebral cortex, striatum, septum, and brainstem. The castration of male mice caused a reduction in levels of β-NGF protein in the hypothalamus and hypophysis, but not in the cerebellum and olfactory bulb, to the femgle levels. The concentrations of β-NGF protein in testicular feminization mice were similar to those in female CD-1 mice in all regions. The concentrations of mRNA for β-NGF in the olfactory bulb and hypophysis from males were higher than those from females. By contrast, northern blots showed no remarkable differences in the amounts of brain-derived neurotrophic factor and neurotrophin-3 between the two sexes. Thus, in some regions of the brain, the production of β-NGF appears to be regulated by testosterone, but the regulatory mechanisms do not appear to be simple. Our present results indicate that the binding of testosterone to its receptor is an important step in the regulation of the level of β-NGF in these region.     

Genetic Transformation in Helicobacter pylori

Genetic transformation in Helicobacter pylori was investigated by using its chromosomal and plasmid DNAs. Six out of the eight strains exhibited the natural competence for incorporation of H. pylori chromosomal DNA, and all the strains incorporated the donor DNA efficiently by washing and concentrating the cells, with a glycerol solution. The much higher frequency of transformation was obtained in each strain by means of electroporation. Electroporation experiments were also conducted by use of the recombinant DNAs consisting of the H. pylori and Escherichia coli plasmids as the donors, and the occurrence of the homologous recombination was demonstrated between the incoming H. pylori plasmid-derived region and the corresponding region of the originally residing plasmid in H. pylori.