(1/840) Intraerythrocytic Plasmodium falciparum expresses a high affinity facilitative hexose transporter.
Asexual stages of Plasmodium falciparum cause severe malaria and are dependent upon host glucose for energy. We have identified a glucose transporter of P. falciparum (PfHT1) and studied its function and expression during parasite development in vitro. PfHT1 is a saturable, sodium-independent, and stereospecific transporter, which is inhibited by cytochalasin B, and has a relatively high affinity for glucose (Km = 0.48 mM) when expressed in Xenopus laevis oocytes. Competition experiments with glucose analogues show that hydroxyl groups at positions C-3 and C-4 are important for ligand binding. mRNA levels for PfHT1, assessed by the quantitative technique of tandem competitive polymerase chain reaction, are highest during the small ring stages of infection and lowest in gametocytes. Confocal immunofluorescence microscopy localizes PfHT1 to the region of the parasite plasma membrane and not to host structures. These findings have implications for development of new drug targets in malaria as well as for understanding of the pathophysiology of severe infection. When hypoglycemia complicates malaria, modeling studies suggest that the high affinity of PfHT1 is likely to increase the relative proportion of glucose taken up by parasites and thereby worsen the clinical condition. (+info)
(2/840) Effect of dietary taurine supplementation on GSH and NAD(P)-redox status, lipid peroxidation, and energy metabolism in diabetic precataractous lens.
PURPOSE: To evaluate changes in glutathione and NAD(P)-redox status, taurine and malondialdehyde (MDA) levels, glucose utilization, and energy metabolism in diabetic precataractous lenses and to assess whether these changes can be prevented with dietary taurine supplementation. METHODS: The experimental groups included control and streptozotocin-diabetic rats with a 3-week duration of diabetes fed unsupplemented or taurine (1% or 5%)-supplemented diets. The levels of glucose, sorbitol, fructose, myo-inositol, oxidized glutathione (GSSG), glycolytic intermediates, malate, alpha-glycerophosphate, and adenine nucleotides were assayed in individual lenses spectrofluorometrically by enzymatic methods, reduced glutathione (GSH) spectrofluorometrically with O-phthaldialdehyde, MDA colorimetrically with N-methyl-2-phenylindole, and taurine by high-performance liquid chromatography. Free cytosolic NAD+/NADH and NADP+/NADPH ratios were calculated from the lactate dehydrogenase and malic enzyme systems. RESULTS: Sorbitol pathway metabolites and MDA were increased, and GSH and taurine levels were reduced in diabetic rats versus controls. The profile of glycolytic intermediates (an increase in glucose 6-phosphate, no change in fructose 6-phosphate and fructose 1,6-diphosphate, an increase in dihydroxyacetone phosphate, a decrease in 3-phosphoglycerate, phosphoenolpyruvate, and pyruvate, and no change in lactate), and a 9.2-fold increase in alpha-glycerophosphate suggest diabetes-induced inhibition of glycolysis. Free cytosolic NAD+/NADH ratios, ATP levels, ATP/ADP, and adenylate charge were reduced, whereas free cytosolic NADP+/NADPH ratios were elevated. Lens taurine levels in diabetic rats were not affected by supplementation with 1% taurine. With 5% taurine supplementation, they were increased approximately 2.2-fold higher than those in untreated diabetics but remained 3.4-fold lower than in controls. Lens GSH levels were similar in diabetic rats fed unsupplemented and 5% taurine-supplemented diets, whereas GSSG and MDA levels and GSSG/GSH ratios were reduced by 5% taurine supplementation. The decrease in free cytosolic NAD+/NADH, ATP/ADP, and adenylate energy charge were ameliorated by 5% taurine supplementation, whereas accumulation of sorbitol pathway intermediates, depletion of myoinositol, inhibition of glycolysis, a decrease in ATP and total adenine nucleotide, and an increase in free cytosolic NADP+/NADPH were not prevented. CONCLUSIONS: Dietary taurine supplementation ameliorates MDA levels, GSSG/GSH, and NAD+/NADH and fails to prevent the osmotically mediated depletion of GSH and taurine and the decrease in glucose utilization and ATP levels in diabetic precataractous lens. Dietary taurine supplementation cannot be regarded as an alternative to aldose reductase inhibition in eliminating antioxidant and metabolic deficits contributing to diabetes-associated cataractogenesis. (+info)
(3/840) Polyol formation and NADPH-dependent reductases in dog retinal capillary pericytes and endothelial cells.
PURPOSE: Dogs fed a diet containing 30% galactose experience retinal vascular changes similar to those in human diabetic retinopathy, with selective pericyte loss as an initial lesion. In the present study the relationship among reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reductases, polyol formation, and flux through the polyol pathway in cultured dog retinal capillary cells were investigated. METHODS: Pericytes and endothelial cells were cultured from retina of beagle dogs. NADPH-dependent reductases were characterized by chromatofocusing after gel filtration. Sugars in cultured cells were analyzed by gas chromatography, and flux through the polyol pathway was investigated by 19F nuclear magnetic resonance (NMR) with 3-fluoro-3-deoxy-D-glucose (3FG) as a substrate. The presence of aldose reductase and sorbitol dehydrogenase in these cells was examined by northern blot analysis. RESULTS: Two distinct peaks corresponding to aldose reductase and aldehyde reductase, the latter being dominant, were observed in pericytes by chromatofocusing. Culture in medium containing either 10 mM D-galactose or 30 mM D-glucose resulted in the accumulation of sugar alcohol in pericytes that was markedly reduced by aldose reductase inhibitors. 19F NMR spectra obtained from pericytes cultured for 5 days in medium containing 2 mM 3FG displayed the marked accumulation of 3-fluoro-deoxysorbitol but not 3-fluoro-deoxyfructose. No 3FG metabolism was observed in similarly cultured endothelial cells. With northern blot analysis, aldose reductase was detected in pericytes but not in endothelial cells. Sorbitol dehydrogenase was below the detectable limit in pericytes and endothelial cells. CONCLUSIONS: Aldose, aldehyde, and glyceraldehyde reductases are present in dog retinal capillary pericytes, with aldehyde reductase being the major reductase present. Polyol accumulation easily occurs in pericytes but not in endothelial cells. (+info)
(4/840) Carbon assimilation by Claviceps purpurea growing as a parasite.
Carbon assimilation by Claviceps purpurea, growing as a parasite on cereals, has been investigated by supplying the host plant with 14CO2 in a closed system. The presence of the pathogen induced the plant to exude photosynthate which contained high levels of sucrose. During the period of 14CO2 supply, 14C was incorporated into the sucrose and so the path of carbon into the parasite could be traced. Hexoses, derived by the action of the fungal sucrase on sucrose, were assimilated by the pathogen and largely converted into polyols - mainly mannitol and, to a lesser extent, trehalose. The rate of carbohydrate metabolism decreased with maturation of the ergot, and also showed qualitative differences between the basal and apical regions of the ergot which were probably a function of nutrient supply. (+info)
(5/840) Involvement of the Escherichia coli phosphoenolpyruvate-dependent phosphotransferase system in regulation of transcription of catabolic genes.
Synthesis of catabolite-sensitive enzymes is repressed in mutants defective in the general proteins (enzyme I and HPr) of the Escherichia coli phosphoenolpyruvate-dependent phosphotransferase system (ptsI and ptsH mutations). To elucidate the mechanism of this phenomenon we constructed isogenic strains carrying pts mutations as well as different lesions of regulation of the lac operon or mutations affecting adenylate cyclase activity (cya mutation) and synthesis of cyclic AMP-receptor protein (crp mutation) Measurements of the differential rate of beta-galactosidase synthesis in these strains showed that the repressive effect of pts mutations was revealed in lac+, lacI, lacOc and cya bacteria, but it was lost in lacP and crp strains. It was concluded that mutational damage to the general components of the phosphoenolpyruvate-dependent phosphotransferase system diminishes activity of the lac promoter. The results obtained led to the conclusion that pts gene products (apparently phospho approximately HPr) are necessary for the initiation of transcription of catabolite-sensitive operons in E. coli. (+info)
(6/840) Sugars and sugar derivatives which inhibit the short-circuit current of the everted small intestine of the rat.
1. The short-circuit current of everted rat intestine supported on a perforated cannula proved to be stable for up to 3 hr and has been used to study competition between transportable and non-transportable sugars. 2. 4,6-O-Ethylidene-alpha-D-glucopyranose (ethylidene glucose) and 4,6-O-benzylidene-e alpha-D-glucopyranos (benzylinene glucose), two nontransportable inhibitors of the hexose transfer system in human erythrocytes, were found to reduce the short-circuit current generated by transportable sugars such as galactose or 3-O-methyl glucose. 3. These compounds were also found to reduce the basal short-circuit current established by the everted intestine in a sugar-free Krebs solution. Both types of inhibition approached saturation at the higher concentrations used. 4. Similar inhibitory properties were shown by mannose, a non-actively accumulated monosaccharide, and by the beta-disaccharides lactose and cellobiose. 5. It is suggested that this common pattern of behaviour is due to the ability of these compounds to react with the sites for active hexose transfer but without translocation by the system. The significance of the inhibition of the basal short-circuit current is briefly discussed in this context. (+info)
(7/840) Structure of the O-specific polysaccharide of a serologically separate strain Proteus penneri 2 from a new proposed serogroup O66.
O-specific polysaccharide chain of Proteus penneri strain 2 lipopolysaccharide was studied by full and partial acid hydrolysis, Smith degradation, methylation analysis, and NMR spectroscopy, including two-dimensional rotating-frame NOE spectroscopy (ROESY) and 1H,13C heteronuclear multiple-quantum coherence (HMQC) experiments. Together with D-glucose and 2-acetamido-2-deoxy-D-glucose, the polysaccharide was found to contain two rarely occurring sugars, 6-deoxy-L-talose (L-6dTal) and 2,3-diacetamido-2,3,6-trideoxy-L-mannose (L-RhaNAc3NAc), and the following structure of a non-stoichiometrically O-acetylated tetrasaccharide repeating unit was established: [equation: see text] The O-specific polysaccharide studied has a unique composition and structure and, accordingly, P. penneri 2 is serologically separate among Proteus strains. Therefore, we propose for P. penneri 2 a new Proteus O-serogroup O66 where this strain is at present the single representative. (+info)
(8/840) Structure of the glucan-binding sugar chain of Tip1p, a cell wall protein of Saccharomyces cerevisiae.
Tip1p is one of the major cell wall mannoproteins of Saccharomyces cerevisiae and is presumed to be synthesized as a glycosylphosphatidylinositol (GPI)-anchored form. We purified Tip1p from a glucanase extract of yeast cell walls and analyzed the sugar chain involved in the cell wall linkage. One mol of glucanase-extracted Tip1p contained 7.5 mol of glucose derived from glucan and 1 mol of ethanolamine, a component of the GPI anchor. One mol of the C-terminal peptide of Tip1p digested with Achromobacter protease I also contained 7.9 mol of glucose and 1 mol of ethanolamine. On the other hand, Tip1p contained no glucosamine, which is a component of the GPI anchor. The glucan-binding sugar chain of Tip1p was released by hydrazinolysis and isolated. This sugar chain contained ethanolamine with a free amino group and a glucose reducing end, but no mannose reducing end. Phosphodiesterase treatment eliminated the free amino group from this sugar chain, suggesting that a phosphodiester bond exists between the ethanolamine and the glucan remnant. These results indicate (1) the glucan-binding sugar chain of Tip1p is a GPI derivative, and (2) the GPI anchor is cleaved at the glycosyl moiety, and the resultant mannose reducing end is probably used to link Tip1p to cell wall glucan. (+info)