Presence of ornithine decarboxylase antizyme in primary cultured hepatocytes of the frog Xenopus laevis

Ornithine decarboxylase (ODC; EC 4.1.1.17) could be induced in primary cultured hepatocytes of the frog, Xenopus laevis, by a hypotonic treatment. Addition of 10 mM putrescine caused a rapid decay of preinduced ODC after a lag period of 30 min. The putrescine-induced ODC decay was faster than the ODC decay in the presence of cycloheximide. Simultaneous addition of cycloheximide blocked the putrescine-induced acceleration of ODC decay, indicating an involvement of protein synthesis. Addition of putrescine to normal medium caused complete loss of ODC activity in 2 h and then ODC-inhibitory activity appeared and progressively increased. The inhibitory factor was non-dialysable and temperature-sensitive and showed a time-independent and stoichiometric pattern of ODC inhibition. On the basis of these observations the inhibitory factor was identified as ODC antizyme. These results indicated that in frog hepatocytes, like in mammalian cells and tissues, ODC is under negative feedback regulation mediated by antizyme.     

Kinetics of hydroperoxide degradation by NADP-glutathione system in mitochondria

Hydroperoxide decomposition by the NADP-glutathione system in rat liver mitochondria was analyzed. Mitochondria were found to contain high concentrations of the reduced form of glutathione (GSH) (4.32 +/- 0.50 nmol/mg) and NADPH (4.74 +/- 0.64 nmol/mg), and high activities of glutathione peroxidase and reductase. In the initial phase of the reaction, the rate of hydroperoxide decomposition was proportional to both the GSH level and the activity of GSH peroxidase. However, in the later steady state, the step of NADP reduction was rate-limiting, and the overall reaction rate was independent of the initial concentration of GSH, and activities of glutathione peroxidase and reductase. Some GSH was released from mitochondria during incubation, but the rate of the decomposition could be simply expressed as kappa [GSH]/2, where kappa is the first-order rate constant of the peroxidase and [GSH] is the intramitochondrial level of GSH in the steady state. The rate of the reaction in the steady state was also dependent on the NADPH level, its reciprocal being linearly correlated with [NADPH]-1. The rate of decomposition of hydroperoxide was influenced by the respiratory state. During state 3 respiration, the rate was greatly depressed, but was still considered to exceed by far the rate of physiological generation of hydroperoxide.     

Length of polymorphisms of restriction fragments of rat mitochondrial DNAs

Differences were found in the HpaII cleavage patterns of two types of rat (Rattus norvegicus) mtDNA, types A and B. One HpaII fragment, Hpa5, of type A was about 30 base pairs smaller than that of type B, but no 30-base pair fragment was detectable in an HpaII digest of type A mtDNA. Moreover, one HaeIII fragment, which is overlapped by Hpa5 in the cleavage map, was also about 30-base pairs smaller in type A than in type B. Thus, the length polymorphism of Hpa5 in the two types is probably not caused by HpaII site gain or loss, but by sequence deletion or insertion.     

Beta-galactosidase-neuraminidase deficiency: restoration of beta-galactosidase activity by protease inhibitors

Beta-Galactosidase was partially restored by protease inhibitors, leupeptin, chymostatin and E-64 in cultured fibroblasts from three patients with beta-galactosidase-neuraminidase deficiency. Pepstatin did not activate this enzyme. Neuraminidase was not affected by any of these compounds in the culture medium. It was concluded that the activating effect was produced by a specific inhibition of thiol proteases.     

Temperature dependent ionic structure of phospholipid monolayers

Radiotracer studies of calcium adsorption to dipalmitoylphosphatidyl-alkanolamine monolayers measured at various temperatures showed that the binding constant of calcium increased with temperature up to around 30°C but then decreased on exceeding this critical temperature.The temperature dependent ionic structure of ampholytic phospholipid monolayers are discussed.     

Desensitization by covalent modification of the chemoreceptor of Escherichia coli

Chemoreceptors in Escherichia coli were studied in situ in chemotactic mutants, deficient in the ability to modify the receptors, by using membrane vesicles prepared from the mutants. The affinity of the receptors for the ligands is related to the level of modification of the receptors. Unmodified serine receptor had a dissociation constant of 0.8 microM, while modified receptor had a dissociation constant that was at least 100-times higher. The results are discussed in relation to the two-state model of the chemoreceptor.     

Oxygen evolution in dark-developed spruce chloroplasts

Chloroplasts isolated from dark-grown seedlings of Picea abiesshowed activities of DCIP and Fecy photoreductions widiout additionof an electron donor to PS-II. In addition to this, when light-inducedoxygen evolution was measured with an oxygen electrode, a significantamount of oxygen was found. These results indicate that thephotoreductions are coupled to the oxygen-evolving reaction.Furthermore, thylakoid membranes were functional in the protonuptake and the 515-nm absorbance change as parameters of theirphysicochemical functions. Electron microscopy showed that thylakoidswere well-developed with prolamellar bodies and partially stackedto from grana. We conclude that oxygen-evolving ability and the physicochemicalfunction of thylakoid membranes develop in chloroplasts of dark-grownspruce seedlings. (Received September 21, 1974; )