2-Deoxy-D-galactose metabolism in ascites hepatoma cells results in phosphate trapping and glycolysis inhibition.

The metabolism of 2-deoxy-D-galactose has been studied in AS-30D rat ascites hepatoma cells in suspension. Using 2-deoxy-D-(1-14C)galactose and an alkaline ethanol deproteinization procedure, the quantitatively identified metabolites included 2-deoxy-D-galactose 1-phosphate comprising 99.3%, and UDP-2-deoxy-D-galactose and UDP-2-deoxy-D-glucose, together amounting to 0.4% of the total metabolites. After incubation for 5 h in the presence of 2-deoxy-D-galactose (1 mmo1/1), the content of 2-deoxy-D-galactose 1-phosphate reached 35 mmo1x(kg cells)-1. The rate of phosphorylation of 2-deoxy-D-galactose was rapid during the first 30 min and decreased to approximately 20% of this rate during the subsequent hours. The rapid trapping of Pi in the form of 2-deoxy-D-galactose 1-phosphate resulted in a depression of free intracellular Pi in spite of a concomitant increase in net 32Pi uptake from the medium and a decrease of ATP and other 5'-nucleotides. The rates of glucose utilization and lactate production were depressed by more than 80% in the presence of 2-deoxy-D-galactose (1 mmo1/1). Interruption of Pi trapping by removal of 2-deoxy-D-galactose from the medium reversed the depressions of Pi and ATP and resulted in a rapid but incomplete relief of glycolysis inhibition. Crossover analysis of glycolytic intermediates indicated an inhibition at the 6-phosphofructokinase step. The depression of glucose utilization may be mediated by the increased level of glucose 6-phosphate, a potent inhibitor of hexokinase. An additional inhibitory effect of a metabolite of 2-deoxy-D-galactose at the 6-phosphofructokinase step was indicated by crossover analysis after reversal of Pi and ATP depressions in the presence of a high intracellular content of 2-deoxy-D-glactose 1-phosphate. The quantitative analysis of the metabolites of 2-deoxy-D-galactose demonstrated the predominance of the monophosphate and the negligible formation of UPD derivatives of this sugar analog in AS-30D hepatoma cells. This provides a system for the investigation of a galactose analog as a phosphate-trapping agent in the virtual absence of uridylate trapping.     

Tissue factor expression in newt iris coincides with thrombin activation and lens regeneration

Lens regeneration in adult salamanders occurs at the pupillary margin of the mid-dorsal iris where pigmented epithelial cells (PEC) re-enter the cell cycle and transdifferentiate into lens. It is not understood how the injury caused by removal of the lens (lentectomy) in one location is linked to initiating the response in a different spatial location (dorsal iris) and to this particular sector. We propose that the blood provides a link between the localised coagulation and signal transduction pathways that lead to regeneration. A transmembrane protein (tissue factor) is expressed in a striking patch-like domain in the dorsal iris of the newt that localises coagulation specifically to this location, but is not expressed in the axolotl, a related species that does not show thrombin activation after lentectomy and cannot regenerate its lens. Our hypothesis is that tissue factor expression localises the initiation of regeneration through the activation of thrombin and the recruitment of blood cells, leading to local growth factor release. This is the first example of gene expression in a patch of cells that prefigures the location of a regenerative response, and links the immune system with the initiation of a regenerative program.     

Effect of retinoic acid on the aggregation of erythrocytes and leukocytes in the blood

Human and animal blood smear staining with PAPh has revealed mononuclear leukocyte-erythrocyte aggregates. Administration of retinoic acid increased concentration and dimensions of these aggregates and was followed by preferential accumulation of PAPh-negative osmotically unstable erythrocytes. Similar changes were detected in the blood samples of women engaged in the production of retinoic acid. Aggregate concentration showed positive correlation with erythrocyte sedimentation rate and no correlation with prothrombin time.     

Fatty acid composition of Escherichia coli cells and their viability in air

Experiments on E. coli used as a model have revealed that fatty-acid composition is one of the characteristics which determine the viability of bacteria in the air. The viability of microbial cells in the air has been shown to increase with the increase of the pool of cyclopropane acids and the palmitic acid/palmitoleic acid ratio in the cells, irrespective of their genotype and the phase of their growth.     

Hexameric purine nucleoside phosphorylase II from Escherichia coli K-12. Physico-chemical and catalytic properties and stabilization with substrates

Some properties of hexameric purine nucleoside phosphorylase II (EC 2.4.2.1) from Escherichia coli K-12 were studied. The enzyme obeys the Michaelis-Menten kinetics with respect to purine substrates (Km for inosine, deoxyinosine and hypoxanthine are equal to 492, 106 and 26.6 microM, respectively) and exhibits negative kinetic cooperativity towards phosphate and ribose-1-phosphate. The Hill coefficient is equal to approximately 0.5 for both substrates. Hexameric purine nucleoside phosphorylase II is not a metal-dependent enzyme; its activity is inhibited by Cu2+, Zn2+, Ni2+ and SO4(2-). The enzyme is the most stable at pH 6.0; it contains essential thiol groups. All substrates partly protect the enzyme against inactivation by 5.5'-dithiobis(2-nitrobenzoic acid) and heat-inactivation and, with the exception of phosphate-against inactivation by p-chloromercuribenzoate. Hypoxanthine, especially in combination with phosphate, afford the best protection against inactivation.     

Die Wuchsstoffleitung in der Pflanze I

Zusammenfassung In Blättern vonColeus-rot und in Kotyledonen vonCucumis sativa wird künstlich durch die Epidermis zugeführter Wuchsstoff (Heteroauxin) apikalwärts geleitet, wenn der direkte Abfluß nach der Mittelrippe unterbunden ist. Daraus folgt, daß die Wuchsstoffleitung in der Pflanze keine streng polare Erscheinung ist, genau so wenig, wie der Transpirationsstrom oder der Strom der Assimilate streng polar fließen.Mit 11 Textabbildungen (16 Einzelbildern).Mit Unterstützung der Deutschen Forschungsgemeinschaft und der William G. Kerckhoff-Stiftung zu Bad Nauheim.     

An improved glucoseoxidase--peroxidase-coupled assay for -fructofuranosidase activity

An improved glucoseoxidase-peroxidase-coupled assay for the determination of β-fructofuranosidase activity is described. The method makes use of the double effect of Tris (2-amino-2-hydroxymethylpropane-1,3-diol) as an inhibitor of both invertase and contaminating glucosidases. The method is very sensitive and is suitable for routine determinations. The total time needed for a single analysis is less than half an hour.