Mercury and the activity of erythrocyte and bone marrow glutathione reductase (E.C. 1.6.4.2.) and glucose-6-phosphate dehydrogenase (E.C. 1.1.1.49) in rats

These studies were carried out on male Wistar rats. Erythrocyte and bone marrow activity of glutathione reductase (GR - E.C. 1.6.4.2) and glucose-6-phosphate dehydrogenase (G6PD - E.C. 1.1.1.49) were determined as affected by mercury. Under physiological conditions the above enzymes were found to be markedly more active in the bone marrow than in erythrocytes. The present experiments have revealed that within the first days there will be an increase in enzyme activity in the bone marrow as well as in glutathione reductase in erythrocytes. Simultaneously, a decline in the activity of erythrocytes could be observed which began within the very first days of the experiments as well as a decrease in the bone marrow, which occurred later in the course of the experiment.     

Studies on the effect of low-molecule uremic toxins on the activity of glucose-6-phosphate dehydrogenase (E.C.1.1.1.49) and transketolase (E.C.2.2.1.1) in human red blood cells

The effect in vitro was studied of methylguanidine (MG) and guanidinosuccinic acid (GSA) on the activity of glucose-6-phosphate dehydrogenase (G-6-P DH) and transketolase (ETKA) of normal erythrocytes. The results show that MG in concentration (1.8 X 10(-5) mol/l) approximate to or higher then its concentration in plasma of patients with chronic renal failure (ch.r.f.) inhibits the activity of G-6-P DH. The changes of similar nature have been observed in case of GSA. In the same incubation conditions MG or GSA bring about the inhibition of ETKA activity in red blood cells. MG and GSA jointly introduced into incubation mixture in concentrations approximate to those appearing in plasma of patients with ch.r.f. cause the inhibition of G-6-P DH by 35.6% on the average, with no statistically significant differences in ETKA activity.     

Kinetic properties of normal human erythrocyte glucose-6-phosphate dehydrogenase dimers.

The steady-state kinetics of normal human erythrocyte glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate: NADP+ oxidoreductase, EC 1.1.1.49) dimers were studied as a function of pH and temperature. Inhibition studies using glucosamine 6-phosphate, NADPH and p-hydroxymercuribenzoate (P-OHMB) were also carried out at pH 8.0. The existence of two binding sites on the enzyme with a transition from low to high affinity for NADP+ when NADP+ concentration is increased is indicated by the nonlinear Lineweaver-Burk plots and sigmoid kinetic patterns. NADPH inhibition was found to be competitive with respect to NADP+ and non-competitive with respect to glucose-6-phosphate. Logarithmic plot of Vmax against pH and inactivation by P-OHMB indicate the participation in the reaction mechanism of imidazolium group of histidine and sulhydryl groups. The initial velocity and product inhibition data gave results which are consistent with the dimeric enzyme following an ordered sequential mechanism. A possible random mechanism is ruled out by the inhibition results of glucosamine 6-phosphate.     

Effects of acetylphenylhydrazine (APHZ) on the stromal cells of the bone marrow in rats (in vitro)

The study has been carried out on Wistar rats. The animals were divided into two groups: the control rats received 0.9% saline solution intraperitoneally in there consecutive days. The experimental animals were administered 1% solution of acetylphenylhydrazine (APHZ) intraperitoneally also in three days. Prior to the experiments samples of blood have been collected from tail veins and reticulocyte counts made: the counts had also been determined before the culture of tissue was started. On the fifth day of the primary culture, the morphology and enzymatic activity of the stromal cells of the bone marrow were studied. The cells grown in vitro were cultured by the use of the Eagle's medium enriched with 20% calf serum and L-glutamate. The type of the cells, their number and rate of growth were noted. In the cells the activity of specific esterase as well as acid and alkaline phosphatase was determined. Statistically significant differences between the control and experimental animals were found both in the numbers of stromal cells and in the differences in score values of activity of particular enzymes in various types of cells.     

Combined erythrocyte glucosephosphate isomerase (GPI) and glucose-6-phosphate dehydrogenase (G6PD) deficiency in an italian family

Summary A severe hemolytic crisis was observed in a 5-yearold boy of Italian origin. Analysis of his hemolysate revealed a hemizygous deficiency of glucose-6-phosphate dehydrogenese (G6PD) and a heterozygous deficiency of glucosephosphate isomerase (GPI). According to the literature this is the fourth family with a combined deficiency of these two enzymes located on different chromosomes. Only the G6PD deficiency seems to be responsible for the hemolytic crisis.Dedicated to Prof. Dr. Walter Sandritter on occasion of his 60th birthday     

The structural subunit of glucose-6-phosphate dehydrogenase (baker's yeast).

The subunit molecular weight of glucose-6-phosphate dehydrogenase (G6PD) from baker's yeast has been evaluated. The subunit molecular weight value is shown to be 25,500 daltons by analytical ultracentrifugation, SDS-polyacrylamide gel electrophoresis, and the number of peptides produced by CNBr cleavage. The number of NADP binding sites was determined to be one per 25,500 dalton unit.     

Specific activities of 6-phosphogluconate dehydrogenase,glucose-6-phosphate dehydrogenase and glucose-6-phosphate isomerase during Bufo bufo development

It has been suggested by some authors that during amphibian development, due to the higher glucose-6-phosphate dehydrogenase (EC 1.1.1.49) activity compared to that of 6-phosphogluconate dehydrogenase (EC 1.1.1.43), 6-phosphogluconate could accumulate in the embryo tissues and regulate the channelling of glucose-6-phosphate into glycolysis. Here, on the base of the specific activities of glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and glucose-6-phosphate isomerase (EC 5.3.1.9) found in the embryos of Bufo bufo during development, it is discussed whether 6-phosphogluconate can accumulate and play a regulative role on glucose-6-phosphate metabolism in the anuran embryo.     

2-deoxy-glucose-6-phosphate utilization in the study of glucose-6-phosphate dehydrogenase mosaicism.

The electrophoretic difference between normal glucose-6-phosphate dehydrogenase (G6PD) and two common variants (G6PD A and G6PD A-) has made the G6PD enzyme system very useful for genetic studies and for investigation on the clonal origin of tumors. This approach has not been possible for another common variant, G6PD mediterranean, which has a normal electrophoretic pattern. The different utilization of 2-deoxy-glucose-6-phosphate (2dG6P), an analog of the normal substrate, by the normal enzyme and the Mediterranean variant, allows a convenient determination of the degree of mosaicism in mononuclear cells from heterozygotes.     

Sublethal ammonia and urea concentrations inhibit rainbow trout (Oncorhynchus mykiss) erythrocyte glucose-6-phosphate dehydrogenase

In vitro and in vivo effects of sublethal ammonia and urea concentrations were assayed on glucose-6-phosphate dehydrogenase (G6PD) of rainbow trout (Oncorhynchus mykiss) erythrocyte. G6PD was purified from erythrocytes with a specific activity of 16.7 EU (mmol NADP+/min)/mg protein and approximately 1600-fold in a yield of approximately 60% by ammonium sulphate precipitation and 2',5'-ADP Sepharose 4B affinity chromatography. The purity of the enzyme was confirmed using SDS polyacrylamide gel electrophoresis. Experiments with ammonia (2.2-5.5 microM) and urea (20-50 microM) showed the inhibitory effects on the enzyme, in vitro. Inhibition effects were determined in vitro by Lineweaver-Burk and regression graphs. The dissociation constant of the enzyme inhibitor complex (Ki) and 50% inhibitory values were 2.26+/-1.21 and 2.86+/-3.51 microM for ammonia and 18.69+/-6.75 and 23.77+/-4.58 microM for urea, respectively. In vivo studies in rainbow trout erythrocytes showed significant (p < 0.01) inhibition of G6PD by ammonia and urea. However, ammonia inhibited more than urea since there were significant differences between the final values of erythrocyte G6PD activities.