Glutathione peroxidase (GPX1) was the first identified selenium-dependent enzyme, and this enzyme has been most useful as a biochemical indicator of selenium (Se) status and the parameter of choice for determining Se ...Glutathione peroxidase (GPX1) was the first identified selenium-dependent enzyme, and this enzyme has been most useful as a biochemical indicator of selenium (Se) status and the parameter of choice for determining Se requirements. We have continued to study Se regulation of GPX1 to better understand the underlying mechanism and to gain insight into how cells themselves regulate nutrient status. In progressive Se deficiency in rats, GPX1 activity,protein and mRNA all decrease in a dramatic, coordinated and exponential fashion such that Se-deficient GPX1 mRNA levels are 6-15% of Sexadequate levels. mRNA levels for other Sedependent proteins are far less decreased in the same animals. The mRNA levels for a second Se-dependent peroxidase, phospholipid hydroperoxide glutathione peroxidase (GPX4 ), are little affected by Se deficiency, demonstrating that Se regulation of GPX1 is unique. Se regulation of GPX1 activity in growing male and female rats shows that the Se requirernent is 100 ng/g diet, based on liver GPX1 activity; use of GPX1 mRNA as the parameter indicates that the Se requirement is nearer to 50 ng Se/g diet in both male and female rats. This approach will readily detect an altered dietary Se requirement, as shown by the incremental increases in dietary Se requirement by 150, 100 or 50 ng Se/g diet in Seudeficient rat pups repleted with Se for 3, 7 or 14 d, respectively. Studies with CHO cells stably transfected with recombinant GPX1 also show that overexpression of GPX1 does not alter the minimum level of media Se necessary for Se-adequate levels of GPX1 activity or mRNA. We hypothesize that classical GPX1 has an integral biological role in the mechanism used by cells to regulate Se status,making GPX1 an especially useful and effective parameter for determining Se requirements in animals展开更多
In the blood fluke Schistosoma mansoni a functionally active, monomeric, phospholipid hydroperoxide glutathione peroxidase (PHGPx) has been purified and characterized. This enzyme contains a catalytically active selen...In the blood fluke Schistosoma mansoni a functionally active, monomeric, phospholipid hydroperoxide glutathione peroxidase (PHGPx) has been purified and characterized. This enzyme contains a catalytically active selenocysteine. The protein has been shown to be the product of a cloned gene, previously referred to as a glutathione peroxidase gene. S. mansoni PHGPx has been found 5 times more abundant in female than in male worm extract. As in vertebrate PHGPx, homology alignment indicates that the residues involved in the glutathione binding by the tetrameric cellular glutathione peroxidase are mutated in the S. mansoni enzyme. Thus, this aspect appears a landmark of the PHGPx-type of glutathione peroxidases,which might be of functional relevance展开更多
A sensitive, micro-5, 5' -dithiobis-2 -nitrobenzise acid (DTNB) assay for determination of glutathione peroxidase (GSH-Px) in mouse blood was reported, Trichloroacetic acid (TCA) 0.16 mol/L was selected as a prote...A sensitive, micro-5, 5' -dithiobis-2 -nitrobenzise acid (DTNB) assay for determination of glutathione peroxidase (GSH-Px) in mouse blood was reported, Trichloroacetic acid (TCA) 0.16 mol/L was selected as a protein pricipitation reagent instead of metaphosphoric acid to remove protein in 10 micro liter blood sample.The surplus glutathione (GSH) reacted with DTNB to form a pale yellow color at 37℃ in pH 6.5 solution. There is good linearity between GSH concenlration and absorbance of pale yellow color measured at 423 nm (r=0.9973). Precision studies for both within day and day-to-day at different concenlrations of DTNB provided CV values of less than 5%. Two kinds of selenium (Se) rich yeast were given to mice (500μg/kg) once a day for ten days by oral abministration. The activity of GSH-Px was measured according to direct DTNB assay. There is high significant difference between treated and control groups. The results show that blood GSH-Px level can be used as an indicator of Se status and the capability of scavenging peroxidatives in mammals.展开更多
文摘Glutathione peroxidase (GPX1) was the first identified selenium-dependent enzyme, and this enzyme has been most useful as a biochemical indicator of selenium (Se) status and the parameter of choice for determining Se requirements. We have continued to study Se regulation of GPX1 to better understand the underlying mechanism and to gain insight into how cells themselves regulate nutrient status. In progressive Se deficiency in rats, GPX1 activity,protein and mRNA all decrease in a dramatic, coordinated and exponential fashion such that Se-deficient GPX1 mRNA levels are 6-15% of Sexadequate levels. mRNA levels for other Sedependent proteins are far less decreased in the same animals. The mRNA levels for a second Se-dependent peroxidase, phospholipid hydroperoxide glutathione peroxidase (GPX4 ), are little affected by Se deficiency, demonstrating that Se regulation of GPX1 is unique. Se regulation of GPX1 activity in growing male and female rats shows that the Se requirernent is 100 ng/g diet, based on liver GPX1 activity; use of GPX1 mRNA as the parameter indicates that the Se requirement is nearer to 50 ng Se/g diet in both male and female rats. This approach will readily detect an altered dietary Se requirement, as shown by the incremental increases in dietary Se requirement by 150, 100 or 50 ng Se/g diet in Seudeficient rat pups repleted with Se for 3, 7 or 14 d, respectively. Studies with CHO cells stably transfected with recombinant GPX1 also show that overexpression of GPX1 does not alter the minimum level of media Se necessary for Se-adequate levels of GPX1 activity or mRNA. We hypothesize that classical GPX1 has an integral biological role in the mechanism used by cells to regulate Se status,making GPX1 an especially useful and effective parameter for determining Se requirements in animals
文摘In the blood fluke Schistosoma mansoni a functionally active, monomeric, phospholipid hydroperoxide glutathione peroxidase (PHGPx) has been purified and characterized. This enzyme contains a catalytically active selenocysteine. The protein has been shown to be the product of a cloned gene, previously referred to as a glutathione peroxidase gene. S. mansoni PHGPx has been found 5 times more abundant in female than in male worm extract. As in vertebrate PHGPx, homology alignment indicates that the residues involved in the glutathione binding by the tetrameric cellular glutathione peroxidase are mutated in the S. mansoni enzyme. Thus, this aspect appears a landmark of the PHGPx-type of glutathione peroxidases,which might be of functional relevance
文摘A sensitive, micro-5, 5' -dithiobis-2 -nitrobenzise acid (DTNB) assay for determination of glutathione peroxidase (GSH-Px) in mouse blood was reported, Trichloroacetic acid (TCA) 0.16 mol/L was selected as a protein pricipitation reagent instead of metaphosphoric acid to remove protein in 10 micro liter blood sample.The surplus glutathione (GSH) reacted with DTNB to form a pale yellow color at 37℃ in pH 6.5 solution. There is good linearity between GSH concenlration and absorbance of pale yellow color measured at 423 nm (r=0.9973). Precision studies for both within day and day-to-day at different concenlrations of DTNB provided CV values of less than 5%. Two kinds of selenium (Se) rich yeast were given to mice (500μg/kg) once a day for ten days by oral abministration. The activity of GSH-Px was measured according to direct DTNB assay. There is high significant difference between treated and control groups. The results show that blood GSH-Px level can be used as an indicator of Se status and the capability of scavenging peroxidatives in mammals.
