Epididymal carbohydrate metabolism. III. Metabolism of the caput and cauda epididymidis after separation from the testis in the rat.
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After unilateral separation of the rat epididymis from the testis, the metabolism of various substrates in vitro by tissue from the attached and separated caput and cauda epididymidis at 7 and 28 days after surgery was determined by radiorespirometry. Hourly collections of 14-CO2 were made during 5-hr incubations. The patterns of 14-CO2 evolution from glucose indicated that most of the metabolic activity followed the Embden-Meyerhof glycolytic and the Krebs cycle respiration pathways. The alteration of the rate of glycolysis was always greater than that of respiration. In all samples, the metabolism of (2-14C) glucose was approximately equal to that of (6-14C) glucose (G-6)and less than that of (1-14C) glucose (G-1). Pentose cycle activity was indicated in all tissues from the caput and cauda epididymidis by the preferential utilization of G-1 over G-6. At 7 and 28 days after surgery, respectively, the G-1:G-6 ratios of 14-CO2 evolution after incubation for 2 hr were 9.75 and 7.79 for the separated caput, 5.17 and 2.66 for the intact caput, 3.11 and 2.52 for the separated cauda and 3.73 and 2.84 for the attached cauda epididymidis. Although epididymal separation did not effect the metabolism of (U-14C) glucose or (U-14C) fructose, glucose appeared to be a more important epididymal substrate than fructose. (+info)
Antagonistic effect of 3,6-dimethamidodibenzopyriodonium gluconate on lipid peroxidation in cerebral cortical neuronal cultures and rat brains during focal cerebral ischemia reperfusion.
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AIM: To study 3,6-dimethamidodibenzopyriodonium gluconate (I-93) antagonistic effects on lipid peroxidation in cerebral cortical neuronal cultures and rat brains during focal cerebral ischemia-reperfusion. METHODS: Cerebral cortical neurons were cultured and rat focal cerebral ischemia-reperfusion model was established by reversible middle cerebral artery occlusion (MCAO) without craniectomy. The efflux of lactate dehydrogenase (LDH) from neurons, content of malondialdehyde (MDA) in neurons and brain homogenate, activity of superoxide dismutase (SOD) in brain homogenate, and index of cerebral edema as well as brain morphology were investigated. RESULTS: I-93 10-40 mumol.L-1 concentration-dependently inhibited efflux of LDH and elevated levels of MDA induced by addition of H2O2(10 mumol.L-1) in vitro. I-93 0.5 mg.kg-1 improved the cerebral morphology, reduced brain edema, decreased MDA content, and enhanced SOD activity in brain homogenate. CONCLUSION: I-93 protects neurons from H2O2-induced neurotoxicity and ischemia-reperfusion mediated damage by increasing the activity of antioxidant enzymes and suppressing the generation of lipid peroxides. (+info)
A deeper investigation on carbohydrate cycling in Sinorhizobium meliloti.
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Recycling of triose-phosphate and pentose-phosphate was previously reported on glucose in Sinorhizobium meliloti, a polysaccharide-synthesizing bacterium, but the metabolic basis of such processes remained unclear. In this work, we have used (13)C-labelling strategies to demonstrate that carbohydrate cycling in this organism is independent of the gluconate bypass, the alternative pathway for glucose assimilation involving its periplasmic oxidation into gluconate. Furthermore, carbohydrate cycling in S. meliloti is also observed on fructose, making the situation in this bacterium significantly different from that depicted for alginate-synthesizing species. (+info)
Regulation of coremium morphogenesis in Penicillium claviforme.
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Coremia of Penicillium claviforme develop in three stages; primordium formation, elongation, and sporulation. Primordium formation was induced by external nutrients, while starvation initiated the differentiation of primordia into coremia with sporeheads. There is strong evidence that external nutrients are not taken up during this differentiation. Continued sporulation by mature coremia again required an external nutrient supply. (+info)
CFTR modulates programmed cell death by decreasing intracellular pH in Chinese hamster lung fibroblasts.
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To study the potential influence of cystic fibrosis conductance regulator (CFTR) on intracellular pH regulation during apoptosis induction, we used PS120 Chinese hamster lung fibroblasts devoid of the Na(+)/H(+) exchanger (NHE1 isoform) transfected with constructs, allowing the expression of CFTR and/or NHE1. Kinetics of lovastatin-induced apoptosis were measured by orcein staining, double staining with Hoechst-33258, propidium iodide, DNA fragmentation, and annexin V labeling. In PS120 control cells, the percentage of apoptotic cells after 40 h of lovastatin treatment was 23 +/- 3%, whereas in PS120 CFTR-transfected cells, this percentage was 40 +/- 4%. In PS120 NHE1 cells, the transfection with CFTR did not modify the percentage of apoptotic cells after 40 h (control: 19 +/- 3%, n = 8; CFTR: 17 +/- 1%, n = 8), indicating that blocking intracellular acidification by overexpressing the Na(+)/H(+) exchanger inhibited the enhancement of apoptosis induced by CFTR. In all cell lines, the initial pH values were identical (pH = 7.46 +/- 0.04, n = 9), and treatment with lovastatin led to intracellular acidification. However, the pH value after 40 h was lower in PS120 CFTR-transfected cells (pH = 6.85 +/- 0.02, n = 10) than in PS120 cells (pH = 7.15 +/- 0.03, n = 10). To further investigate the origin of this increased intracellular acidification observed in CFTR-transfected cells, the activity of the DIDS-inhibitable Cl(-)/HCO exchanger was studied. 8-Bromoadenosine 3',5'-cyclic monophosphate incubation resulted in Cl(-)/HCO exchanger activation in PS120 CFTR-transfected cells but had no effect on PS120 cells. Together, our results suggest that CFTR can enhance apoptosis in Chinese hamster lung fibroblasts, probably due to the modulation of the Cl(-)/HCO exchanger, resulting in a more efficient intracellular acidification. (+info)
Studies on the subsite structure of amylases. I. Interaction of glucoamylase with substrate and analogues studied by difference-spectrophotometry.
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Studies were made on the ultraviolet difference-spectra of glucoamylase from Rhizopus niveus [EC 3.2.1.3] specifically produced by the substrate maltose and the inhibitors, glucose, glucono-1: 5-lactone (gluconolactone), methyl beta-D-glucoside, cellubiose, and cyclohexa-, and cyclohepta-amyloses. Of these, maltose and gluconolactone produced characteristic difference spectra with a trough near 300 nm. Based on studies with a model compound for a tryptophan residue, Ac-Trp, this trough was attributed to the effect of a negative charge upon the tryptophan residue. From the concentration dependency of the difference spectra, the dissociation constants of the complexes between the enzyme and maltose, glucose, and gluconolactone were evaluated to be 1.2 mM, 51 mM, and 1.5 mM, respectively. These values are in good agreement with the values of Km or K1 obtained from the steady-state kinetics. The difference-spectrophotometric data suggested that referring to the values of subsite affinities of glucoamylase, maltose, and gluconolactone occupy mainly Subsite 1, where the non-reducing-end glucose residue of a substrate is bound in a productive form and that a tryptophan residue with shows a trough near 300 nm in difference spectra is located in this subsite. (+info)
Catabolite repression of the citrate fermentation genes in Klebsiella pneumoniae: evidence for involvement of the cyclic AMP receptor protein.
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Klebsiella pneumoniae is able to grow anaerobically with citrate as a sole carbon and energy source by a fermentative pathway involving the Na(+)-dependent citrate carrier CitS, citrate lyase, and oxaloacetate decarboxylase. The corresponding genes are organized in the divergent citC and citS operons, whose expression is strictly dependent on the citrate-sensing CitA-CitB two-component system. Evidence is provided here that the citrate fermentation genes are subject to catabolite repression, since anaerobic cultivation with a mixture of citrate and glucose or citrate and gluconate resulted in diauxic growth. Glucose, gluconate, and also glycerol decreased the expression of a chromosomal citS-lacZ fusion by 60 to 75%, whereas a direct inhibition of the citrate fermentation enzymes was not observed. The purified cyclic AMP (cAMP) receptor protein (CRP) of K. pneumoniae bound to two sites in the citC-citS intergenic region, which were centered at position -41.5 upstream of the citC and citS transcriptional start sites. Binding was apparently stimulated by the response regulator CitB. These data indicate that catabolite repression of the citrate fermentation genes is exerted by CRP and that in the absence of repressing carbon sources the cAMP-CRP complex serves to enhance the basal, CitB-dependent transcription level. (+info)
Purification and some properties of a beta-glucosidase from Trichoderma harzianum type C-4.
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Type C-4 strain of Trichoderma harzianum was isolated as a microorganism with high cellulolytic activity. Beta-glucosidase is involved in the last step of cellulose saccharification by degrading cellobiose to glucose, and plays an important role in the cellulase enzyme system with a synergic action with endoglucanase and cellobiohydrolase for cellulose degradation. Beta-glucosidase from T. harzianum type C-4 was purified to homogeneity through Sephacryl S-300, DEAE-Sephadex A-50, and Mono P column chromatographies. It was a single polypeptide with the molecular mass of 75,000 by SDS-PAGE. The enzyme was very active at pH 5.0 and 45 degrees C. No significant inhibition was observed in the presence of metal ions, thiol reagents, or EDTA. The enzyme was stable in the presence of 5% ox gall and digestive enzymes. p-Nitrophenyl-beta-D-cellobioside worked as a substrate for the enzyme as much as p-nitrophenyl-beta-glucopyranoside. Glucose and gluconolactone showed competitive inhibition with a Ki of 1 mM and 1.8 microM, respectively, while galactose, mannose, and xylose did not inhibit the enzyme significantly. (+info)