New pteridine substrates for dihydropteridine reductase and horseradish peroxidase.

The oxidation of 4,5-diaminopyrimidin-6(1H)-one, 5,6,7,8-tetrahydropteridin-4(3H)-one, its 6-methyl and cis-6,7-dimethyl derivatives, and 6-methyl- and cis-6-7-dimethyl-5,6,7,8-tetrahydropterins, by horseradish peroxidase/H2O2 is enzymic and follows Michaelis-Menten kinetics, and its Km and kcat. values were determined. This oxidation of 5,6,7,8-tetrahydropterins produces quinonoid dihydropterins of established structure, and they are known to be specific substrates for dihydropteridine reductase. By analogy the peroxidase/H2O2 oxidation of the 5,6,7,8-tetrahydropteridin-4(3H)-ones should produce similar quinonoid dihydro species. The quinonoid species derived from 5,6,7,8-tetrahydropteridin-4(3H)-one and its 6-methyl and cis-6,7-dimethyl derivatives are shown to be viable substrates for human brain dihydropteridine reductase, and apparent Km and Vmax. values are reported.     

A simple method for preparing physiologically active mitochondria from plant leaves rich in oils and phenolics

Amberlite XAD-7, a nonionic polyacrylate adsorbent, was found to be a very effective protectant for isolating mitochondria from tissues rich in oils and phenolics. Physiologically active, well-coupled mitochondria were successfully prepared from young green leaf tissues of citrus, apple, pear and tobacco.Abbreviations DNP 2,4-dinitrophenol - CCCP carbonyl cyanide m-chlorophenylhydrazone - BSA bovine serum albumin - PVP polyvinylpyrrolidone     

Studies on the interactions between phospholipids and membrane-bound enzymes in microsomes. Effects of phospholipases C on the glucose-6-phosphatase system of rat liver microsomes

The role of phospholipids in the glucose-6-phosphatase system, including glucose-6-P phosphohydrolase and glucose-6-P translocase, was studied in rat liver microsomes by using phospholipases C and detergents. In the time course experiments on detergent exposure, the maximal activation of glucose-6-P phosphohydrolase varied according to the nature of the detergent used. On treatment of microsomes with phospholipase C of C. perfringens, the activity of glucose-6-P phosphohydrolase without detergent (i.e. without rupture of translocase activity) was gradually decreased with the progressive hydrolysis of phosphatidylcholine and phosphatidylethanolamine on the microsomal membrane, and was restored by incubation of these microsomes with egg yolk phospholipids. The extent of decrease in this phosphohydrolase activity in the detergent-exposed microsomes (with rupture of translocase activity) also varied depending on the detergent used (Triton X-114 or taurocholate). When 66% of the phosphatidylinositol on the membrane was hydrolyzed by phosphatidylinositol-specific phospholipase C of B. thuringiensis, the inhibition of glucose-6-P phosphohydrolase activity without detergent was very small. Although the inhibition of enzyme activity with detergent was apparently greater than that without detergent, the enzyme activity was stimulated by the breakdown of phosphatidylinositol when the enzyme activity was measured at lower concentration (0.5 mM) of substrate, glucose-6-P. The latency of mannose-6-P phosphohydrolase, a plausible index of microsomal integrity, remained above 70% after the hydrolysis of phosphatidylcholine, phosphatidylethanolamine, or phosphatidylinositol. The results show that the glucose-6-phosphatase system requires microsomal phospholipids for its integrity, suggesting that there exists a close relation between phosphatidylinositol and glucose-6-P translocase.     

Effect of alpha-fluoromethylhistidine on the histamine content of the brain of W/Wv mice devoid of mast cells: turnover of brain histamine

In the brains of W/Wv mutant mice that have no mast cells, the histidine decarboxylase (HDC) level is as high as in the brain of congenic normal mice (+/+), but the histamine content is 53% of that of +/+ mice. The effects of alpha-fluoromethylhistidine (alpha-FMH) on the HDC activity and histamine content of the brain of W/Wv and +/+ mice were examined. In both strains, 30 min after i.p. injection of alpha-FMH the HDC activity of the brain had decreased to 10% of that in untreated mice. The histamine content decreased more gradually, and after 6 h about half of the control level remained in +/+ mice, whereas histamine had disappeared almost completely in W/Wv mice. It is concluded that the portion of the histamine content that was depleted by HDC inhibitor in a short time is derived from non-mast cells, probably neural cells. The half-life of histamine in the brain of W/Wv mice was estimated from the time-dependent decrease in the histamine content of the brain after administration of alpha-FMH: 48 min in the forebrain, 103 min in the midbrain, and 66 min in the hindbrain.     

Power requirement in non-newtonian fermentation broth

Power requirements in the agitation of non-Newtonian fermentation broths with and without aeration were measured by a strain gage-type dynamometer. Broth from the production of gluc-amylase by Endomyces species and carboxymethyl cellulose solutions were used as non-Newtonian fluids. In gas–liquid agitation systems, the correlation between P_g and P₀² ND³/Q^0.56 observed by Michel and Miller was found to be applicable to non-Newtonian fluids in laminar and transition regions. This was particularly true for fluids with apparent viscosities larger than 300 cp. The impeller diameter and impeller blade width had considerable effects on power consumption in a nongassed system. It was suggested, therefore, that P_g/P₀ should be correlated by a dimensionless term involving some impeller-size factors.     

Spatio-temporal distribution patterns of two tachinid flies,Epicampocera succincta andCompsilura concinnata parasitizingPieris

Larvae of genus Pieris in the northern part of Kyoto City are parasitized by two tachinid flies:Epicampocera succincta, a specialist on genus Pieris, and Compsilura concinnata, a generalist with very wide host-range. We surveyed the parasitism rates of Pieris by both flies for two years at six study areas. In these study areas, there lived three host species in the genus Pieris: P. rapae, P. melete, and P. napi, but neither tachinid parasitized P. napi to any significant extent. In the mountainous district, P. rapae and P. melete coexisted and their populations were relatively continuous, while in the lowland, only P. rapae larvae were abundant in spring and autumn, but even they disappeared in summer. Parasitisms by E. succincta occurred mainly in mountainous district and never in the lowland. C. concinnata parasitized Pieris in all the areas, but its parasitisms occurred mainly in autumn. We analyzed the factors affecting the spatial and temporal patterns of parasitism rates and presumed that the temporal discontinuity of host population restricted the distribution of the specialist parasitoid.     

Analysis of the Epitopes Recognized by Mouse Monoclonal Antibodies Directed to Yersinia enterocolitica Heat-Shock Protein 60

To determine amino acid sequences of the epitopes recognized by monoclonal antibodies (mAbs) 3C8 and 5C3 directed against Yersinia enterocolitica heat-shock protein (HSP60), a dot blot analysis was perfomed using synthesized peptides of Y. enterocolitica HSP60 such as peptides p316-342, p327-359, p340-366, p316-326, p316-321, p319-323, and p321-326 which represent positions of amino acids in Y. enterocolitica HSP60. The dot blot analysis revealed that 5C3 mAb reacted with p316-342, p316-326 and p321-326, and 3C8 mAb p316-342 and p316-326. These results indicate that the epitopes recognized by the mAbs were associated with eleven amino acids, Asp Leu Gly Gln Ala Lys Arg Val Val Ile Asn, of p316-326. The sequence homology between p316-326 of Y. enterocolitica HSP60 and the rest of the HSP60 family suggests that the five amino acids of Lys, Arg, Val, Ile and Asn, which are highly conserved in the HSP60 family, might be related with the epitope recognized by 3C8. In contrast, it was also demonstrated that three amino acids of Leu, Gly and Val, which are not well conserved in the HSP60 family, might be related to the epitope recognized by 5C3.     

A temperature-sensitive osmophilic mutant of Zygosaccharomyces rouxii

Summary A temperature-sensitive osmophilic mutant of Zygosaccharomyces rouxii, OS15, was isolated, which required high salt or sugar concentration for growth above 30°C. Cell viability at 35°C in the presence of NaCl was higher than in the absence of NaCl, and a survival ratio of the mutant cells after incubation at 55°C was also higher in the presence of NaCl than NaCl-free condition. Furthermore, resistance to UV light, hygromycin B and geneticin was improved in the presence of NaCl. There was no difference between the parent and the mutant in fatty acid saturation and microscopic cell shape under NaCl condition.