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1.
The B1-antivitamin activity of oxythiamine disulphide monosulphoxide has been determined in experiments on albino mice. It is shown that this derivative is less toxic and exerts a more profound and prolonged inhibitory action on the pyruvate dehydrogenase and transketolase activity in the animal body in comparison with initial oxythiamine disulphide. 相似文献
2.
B1-antivitamin activity of symmetrical oxythiamine disulphide esters with succinic and o-phthalic acids has been studied in experiments on albino mice. It is shown that O-acyloxythiamine disulphides exert more profound and prolonged inhibitory effect on the transketolase activity in the animal body in comparison with the known antagonist of vitamin B1, oxythiamine, and the initial oxythiamine disulphide. 相似文献
3.
The experiments on mice have shown that oxythiamine disulphide derivatives with the branched hydrocarbon chains are less toxic in the organism as compared to oxythiamine and corresponding disulphides with the unbranched hydrocarbon chains and also induce a more pronounced inhibition of transketolase in the liver and other tissues. It is found that under the effect of the above substances the recovery of enzymic activity is slower than in the case with the oxythiamine application. 相似文献
4.
Modification of the method for transketolase purification in the rat liver was used to reveal the existence of two molecular forms of this enzyme. One of the forms does not react to development of avitaminosis in animals caused by oxythiamine in different doses. The method is developed for isolation of the basic transketolase form in the rat liver with the 50-80% yield of activity. Km values of two sites for coenzyme binding on protein do not depend on the extent of holoenzyme reconstruction from apoenzyme. 相似文献
5.
Increasing doses of oxythiamine were studied as exerting the effect on transketolase inactivation in rat tissues. A conclusion is made that in the process of synthesis de novo there is a transient form of the enzyme accessible for interaction with oxythiamine pyrophosphate. Injection of oxythiamine in the increasing doses are accompanied by a decrease in the glycogen amount, increase in the intracellular level most of the studied intermediates of glycolysis and pentose cycle as well as cAMP. The probable biochemical mechanism of the oxythiamine action is connected with the activation of processes dependent on cAMP. 相似文献
6.
Tylicki A Czerniecki J Dobrzyn P Matanowska A Olechno A Strumilo S 《Canadian journal of microbiology》2005,51(10):833-839
Oxythiamine is an antivitamin derivative of thiamine that after phosphorylation to oxythiamine pyro phosphate can bind to the active centres of thiamine-dependent enzymes. In the present study, the effect of oxythiamine on the viability of Saccharomyces cerevisiae and the activity of thiamine pyrophosphate dependent enzymes in yeast cells has been investigated. We observed a decrease in pyruvate decarboxylase specific activity on both a control and an oxythiamine medium after the first 6 h of culture. The cytosolic enzymes transketolase and pyruvate decarboxylase decreased their specific activity in the presence of oxythiamine but only during the beginning of the cultivation. However, after 12 h of cultivation, oxythiamine-treated cells showed higher specific activity of cytosolic enzymes. More over, it was established by SDS-PAGE that the high specific activity of pyruvate decarboxylase was followed by an increase in the amount of the enzyme protein. In contrast, the mitochondrial enzymes, pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase complexes, were inhibited by oxythiamine during the entire experiment. Our results suggest that the observed strong decrease in growth rate and viability of yeast on medium with oxythiamine may be due to stronger inhibition of mitochondrial pyruvate dehydrogenase than of cytosolic enzymes. 相似文献
7.
A number of possible affinity adsorbents for transketolase (sedoheptulose-7-phosphate:D-glyceraldehyde-3-phosphateglycolaldehydetransferase, EC 2.2.1.1) were prepared. The behaviour of the enzyme from Candida utilis and from Baker's yeast on columns of these and of Blue Sepharose CL-6B was examined, together with the behaviour of the contaminating enzyme, ribulose 5-phosphate 3-epimerase (EC 5.1.3.1). A procedure for removing bound thiamine pyrophosphate by dialysis against EDTA was developed. The competitive inhibition of transketolase by oxythiamine and neopyrithiamine was measured and the Ki values obtained of 1.4 and 4.3 mM, respectively, were compared with the affinity of adsorbents prepared from these two inhibitors. Adsorbents containing bound thiamine pyrophosphate were relatively ineffective but those containing epoxy-linked neopyrithiamine and D-ribose 5-phosphate adsorbed the enzyme at pH 7.4 and it could be eluted in a specific manner. 相似文献
8.
Solov'eva ON Meshalkina LE Kovina MV Selivanov VA Bykova IA Kochetov GA 《Biochemistry. Biokhimii?a》2000,65(10):1202-1205
Two substrates of the transketolase reaction are known to bind with the enzyme according to a ping-pong mechanism [1]. It is shown in this work that high concentrations of ribose-5-phosphate (acceptor substrate) compete with xylulose-5-phosphate (donor substrate), suppressing the transketolase activity (Ki = 3.8 mM). However, interacting with the donor-substrate binding site on the protein molecule, the acceptor substrate, unlike the donor substrate, does not cause any change in the active site of the enzyme. The data are interesting in terms of studying the regulatory mechanism of the transketolase activity and the structure of the enzyme-substrate complex. 相似文献
9.
V V Vinogradov K A Mandrik S A Strumilo S K Matsiuk 《Biokhimii?a (Moscow, Russia)》1979,44(5):868-875
Gel filtration and equilibrium dialysis demonstrated that the hyaloplasmic fraction of the liver of B1-deficient rats does not practically bind C-TDP in vitro. An addition of the excess of non-labelled coenzyme does not increase the transketolase activity. The data obtained suggest that transketolase activation in the hyaloplasmic fraction of the liver of B1-deficient rats after administration of thiamine in vivo is due to stimulation of the additional synthesis of the enzyme protein rather than to the saturation of the free apoenzyme with newly-formed TDP. In vivo and in vitro studies suggest that the hyaloplasmic fraction of the liver of B1-deficient rats contains no free apoenzyme of transketolase. 相似文献
10.
SS Chng RJ Dutton K Denoncin D Vertommen JF Collet H Kadokura J Beckwith 《Molecular microbiology》2012,85(5):996-1006
Escherichia coli uses the DsbA/DsbB system for introducing disulphide bonds into proteins in the cell envelope. Deleting either dsbA or dsbB or both reduces disulphide bond formation but does not entirely eliminate it. Whether such background disulphide bond forming activity is enzyme-catalysed is not known. To identify possible cellular factors that might contribute to the background activity, we studied the effects of overexpressing endogenous proteins on disulphide bond formation in the periplasm. We find that overexpressing PspE, a periplasmic rhodanese, partially restores substantial disulphide bond formation to a dsbA strain. This activity depends on DsbC, the bacterial disulphide bond isomerase, but not on DsbB. We show that overexpressed PspE is oxidized to the sulphenic acid form and reacts with substrate proteins to form mixed disulphide adducts. DsbC either prevents the formation of these mixed disulphides or resolves these adducts subsequently. In the process, DsbC itself gets oxidized and proceeds to catalyse disulphide bond formation. Although this PspE/DsbC system is not responsible for the background disulphide bond forming activity, we suggest that it might be utilized in other organisms lacking the DsbA/DsbB system. 相似文献
11.
Human erythrocyte adenyl and pyridine nucleotide production has been tested in cell-free lysates and in intact cells. The main products obtained in cells incubated with adenine and nicotinic acid are adenosine triphosphate and nicotinate mononucleotide, respectively, under any experimental condition used (incubation time, base concentration). Adenine-phosphoribosyltransferase activity determined in crude lysates is about 100 times higher than nicotinate-phosphoribosyltransferase activity, while cellular adenyl nucleotide production is only three times higher than that of pyridine nucleotide. A strong intracellular regulation for the former, but not latter, synthetic process is thus suggested. Intact erythrocyte nicotinate nucleotide production is inhibited by adenine, while nicotinate-phosphoribosyltransferase activity is not. The possible regulation by adenyl nucleotides is discussed in light of the modulating action of ATP on nicotinate-phosphoribosyltransferase activity. The kinetic characteristics of both adenine- and nicotinate-phosphoribosyltransferases, determined on crude lysates, are reported. 相似文献
12.
Thiamine-deficiency in animals induced by everyday subcutaneous administration of oxythiamine in a dose of 4, 40 and 100 mg/kg of weight for 10 days results in a decrease of the total activity of pyruvate kinase in the liver tissue and does not affect the mentioned index in the kidney and heart tissues. It is shown that as a result of the enzyme fractionation in the column with DEAE-cellulose the total activity of pyruvate kinase in the liver tissue of rats with thiamine-deficiency decreases due to L-isoform while the content of M-isoform remains unchanged. Thiamine deficiency does not affect kinetic characteristics of the L-isoform, extracted from the liver and this shows the absence of changes in the degree of phosphorylation of pyruvate kinase L-isoform under these conditions. 相似文献
13.
P P Filippov I K Shestakova N K Tikhomirova G A Kochetov 《Biokhimii?a (Moscow, Russia)》1979,44(3):521-528
Some properties of homogeneous transketolase from pig liver were studied. It was shown that the pH optimum of the transketolase reaction lies within the range of 7.8--8.2. The isoelectric point is at pH 7.6--7.8. The molecular weight of transketolase is 138,000 +/- 3,000 as determined by the sedimentation equilibrium method and about 152,000 according to the data from gel filtration through Sephadex G-200. The enzyme molecule is a tetramer of the alpha 2 beta 2 type. The molecular weights of the alpha- and beta- subunits determined by polyacrylamide gel in the presence of sodium dodecyl sulfate are 52,000--56,000 and 27,000--29,000, respectively. Transketolase contains about two moles of TPP per mole of protein and does not require metal ions for its catalytic activity. 相似文献
14.
The effect of thiamine supplementation on tumour proliferation. A metabolic control analysis study. 总被引:2,自引:0,他引:2
B Comín-Anduix J Boren S Martinez C Moro J J Centelles R Trebukhina N Petushok W N Lee L G Boros M Cascante 《European journal of biochemistry》2001,268(15):4177-4182
Thiamine deficiency frequently occurs in patients with advanced cancer and therefore thiamine supplementation is used as nutritional support. Thiamine (vitamin B1) is metabolized to thiamine pyrophosphate, the cofactor of transketolase, which is involved in ribose synthesis, necessary for cell replication. Thus, it is important to determine whether the benefits of thiamine supplementation outweigh the risks of tumor proliferation. Using oxythiamine (an irreversible inhibitor of transketolase) and metabolic control analysis (MCA) methods, we measured an in vivo tumour growth control coefficient of 0.9 for the thiamine-transketolase complex in mice with Ehrlich's ascites tumour. Thus, transketolase enzyme and thiamine clearly determine cell proliferation in the Ehrlich's ascites tumour model. This high control coefficient allows us to predict that in advanced tumours, which are commonly thiamine deficient, supplementation of thiamine could significantly increase tumour growth through transketolase activation. The effect of thiamine supplementation on tumour proliferation was demonstrated by in vivo experiments in mice with the ascites tumour. Thiamine supplementation in doses between 12.5 and 250 times the recommended dietary allowance (RDA) for mice were administered starting on day four of tumour inoculation. We observed a high stimulatory effect on tumour growth of 164% compared to controls at a thiamine dose of 25 times the RDA. This growth stimulatory effect was predicted on the basis of correction of the pre-existing level of thiamine deficiency (42%), as assayed by the cofactor/enzyme ratio. Interestingly, at very high overdoses of thiamine, approximately 2500 times the RDA, thiamine supplementation had the opposite effect and caused 10% inhibition of tumour growth. This effect was heightened, resulting in a 36% decrease, when thiamine supplementation was administered from the 7th day prior to tumour inoculation. Our results show that thiamine supplementation sufficient to correct existing thiamine deficiency stimulates tumour proliferation as predicted by MCA. The tumour inhibitory effect at high doses of thiamine is unexplained and merits further study. 相似文献
15.
M J Janssen H M Verheij A J Slotboom M R Egmond 《European journal of biochemistry》1999,261(1):197-207
Secretory phospholipases A2 (PLA2s) are small homologous proteins rich in disulphide bridges. These PLA2s have been classified into several groups based on the disulphide bond patterns found [Dennis, E. A. (1997) Trends Biochem. Sci. 22, 1-2]. To probe the effect of the various disulphide bond patterns on folding, stability and enzymatic properties, analogues of the secretory PLA2s were produced by protein engineering of porcine pancreatic PLA2. Refolding experiments indicate that small structural variations play an important role in the folding of newly made PLA2 analogues. Introduction of a C-terminal extension together with disulphide bridge 50-131 gives rise to an enzyme that displays full enzymatic activity having increased conformational stability. In contrast, introduction of a small insertion between positions 88 and 89 together with disulphide bridge 86-89 decreases the catalytic activity significantly, but does not change the stability. Both disulphide bridges 11-77 and 61-91 are important for the kinetic properties and stability of the enzyme. Disulphide bridge 11-77, but not 61-91, was found to be essential to resist tryptic breakdown of native porcine pancreatic PLA2. 相似文献
16.
The effect of hexacyanoferrate(III) on the catalytic activity of transketolase has been studied. This oxidant inactivates
only one of two active sites of the enzyme, the one with a higher affinity to the coenzyme (thiamine diphosphate). The second
active site does not lose its catalytic activity. These observations indicate that the active sites of holotransketolase,
being indiscernible by data of X-ray analysis, exhibit functional nonequivalence. 相似文献
17.
M V Jones 《Archives of microbiology》1975,102(3):275-279
The activity of the membrane-bound ascorbate-TMPD oxidase in Pseudomonas putida varies with growth conditions and age of the culture. A comparison of the effects of cyanide and azide on the oxidation of various substrates suggests that ascorbate-TMPD oxidase is not the terminal oxidase for NADH or succinate oxidation. However, it does have a role in the oxidation of nicotinate, and may act as an additional terminal oxidase under certain other growth conditions. 相似文献
18.
M Ali C J Gubler J Al Saleh F Abu Farsak 《Comparative biochemistry and physiology. B, Comparative biochemistry》1987,87(4):833-835
1. A study was made of transketolase activity in red and white blood cells and of conditions for assay for transketolase activity and for assessment of the "TPP effect" in human and rat blood. 2. The ratio of the transketolase activity in white cells to that in red cells varied between 23 and 93. 3. Red cells or white cells can both be used for assessment of transketolase activity and the "TPP effect", but the best source for evaluation of transketolase activity and the percent change on addition of thiamin diphosphate appears to be whole blood. 相似文献
19.
Mechanisms regulating adipose tissue pyruvate dehydrogenase 总被引:21,自引:20,他引:1
1. Isolated rat epididymal fat-cell mitochondria showed an inverse relationship between ATP content and pyruvate dehydrogenase activity consistent with competitive inhibition of pyruvate dehydrogenase kinase by ADP. At constant ATP concentration pyruvate rapidly activated pyruvate dehydrogenase in fat-cell mitochondria, an observation consistent with inhibition of fat-cell pyruvate dehydrogenase kinase by pyruvate. Pyruvate dehydrogenase in fat-cell mitochondria was also activated by nicotinate (100mum) and by extramitochondrial Na(+) (replacing K(+)) but not by ouabain or insulin. 2. In rat epididymal fat-pads incubated in vitro pyruvate dehydrogenase was activated by addition of insulin in the absence of substrate or in the presence of glucose (10mm) or fructose (10mm). Glucose and fructose activated the dehydrogenase in the absence or in the presence of insulin, and pyruvate also activated in the absence of insulin. It is concluded that extracellular glucose, fructose and pyruvate may activate the dehydrogenase by raising intracellular pyruvate and that insulin may activate the dehydrogenase by some other mechanism. 3. Ouabain (300mum) and medium in which K(+) was replaced by Na(+), activated pyruvate dehydrogenase in epididymal fat-pads. Prostaglandin E(1) (1mug/ml), 5-methylpyrazole-3-carboxylate (10mum) and nicotinate (10mum), which are as effective as insulin as inhibitors of lipolysis and which like insulin lower tissue concentration of cyclic AMP (adenosine 3':5'-cyclic monophosphate), did not activate pyruvate dehydrogenase. Higher concentrations of prostaglandin E(1) (10mug/ml) and nicotinate (100mum) produced some activation of the dehydrogenase. 4. It is concluded that the activation of pyruvate dehydrogenase by insulin is not due to the antilipolytic effect of the hormone and that the action of insulin in lowering adipose-cell concentrations of cyclic AMP does not afford an obvious explanation for the effect of the hormone on pyruvate dehydrogenase. The possibility that the effects of insulin, ouabain and K(+)-free medium may be mediated by Ca(2+) is discussed. 相似文献
20.
Gerhard Schenk Ronald G. Duggleby Peter F. Nixon 《The international journal of biochemistry & cell biology》1998,30(12):1297-1318
This review highlights recent research on the properties and functions of the enzyme transketolase, which requires thiamin diphosphate and a divalent metal ion for its activity. The transketolase-catalysed reaction is part of the pentose phosphate pathway, where transketolase appears to control the non-oxidative branch of this pathway, although the overall flux of labelled substrates remains controversial. Yeast transketolase is one of several thiamin diphosphate dependent enzymes whose three-dimensional structures have been determined. Together with mutational analysis these structural data have led to detailed understanding of thiamin diphosphate catalysed reactions. In the homodimer transketolase the two catalytic sites, where dihydroxyethyl groups are transferred from ketose donors to aldose acceptors, are formed at the interface between the two subunits, where the thiazole and pyrimidine rings of thiamin diphosphate are bound. Transketolase is ubiquitous and more than 30 full-length sequences are known. The encoded protein sequences contain two motifs of high homology; one common to all thiamin diphosphate-dependent enzymes and the other a unique transketolase motif. All characterised transketolases have similar kinetic and physical properties, but the mammalian enzymes are more selective in substrate utilisation than the nonmammalian representatives. Since products of the transketolase-catalysed reaction serve as precursors for a number of synthetic compounds this enzyme has been exploited for industrial applications. Putative mutant forms of transketolase, once believed to predispose to disease, have not stood up to scrutiny. However, a modification of transketolase is a marker for Alzheimer’s disease, and transketolase activity in erythrocytes is a measure of thiamin nutrition. The cornea contains a particularly high transketolase concentration, consistent with the proposal that pentose phosphate pathway activity has a role in the removal of light-generated radicals. 相似文献