Thiamine-binding activity of Saccharomyces cerevisiae plasma membrane

The specific binding activity to [14C]thiamine was found to be located in hte plasma membrane of Saccharomyces cerevisiae. The activity was inhibited by several thiamine analogs and it was hardly detectable in the plasma membrane from a thiamine transport mutant of Saccharomyces cerevisiae. Some properties of the thiamine-binding activity of yeast plasma membrane are discussed in connection with those of the thiamine transport system.     

Occurrence of thiaminase II in Saccharomyces cerevisiae

It was found that cell-free extracts of Saccharomyces cerevisiae contain thiaminase II which hydrolyzes thiamine and thiamine analogs. The possible involvement of this enzyme and thiamine-synthesizing enzymes in thiamine production from thiamine antagonists is discussed.     

Reversal of pyrithiamine-induced growth inhibition ofSaccharomyces cerevisiae by oxythiamine

Summary Oxythiamine reversed the growth inhibition ofSaccharomyces cerevisiae caused by pyrithiamine, although oxythiamine alone inhibited yeast cell growth. This phenomenon was explained by thiamine production from these 2 thiamine antagonists which was demonstrated using cell suspensions and the crude extract ofS. cerevisiae.     

Inhibition of thiamine transport in baker's yeast by methylene blue

Summary Methylene blue was found to inhibit thiamine transport competitively (K_i=0.63 M) in baker's yeast. The dye was also effective in abolishing the growth inhibition ofSaccharomyces cerevisiae by pyrithiamine which is known to be taken up by a common transport system for thiamine in yeast cells. A possible mechanism for the inhibition by methylene blue of the thiamine transport system in baker's yeast is discussed.     

Thiamine triphosphate and thiamine triphosphatase activities: from bacteria to mammals

In most organisms, the main form of thiamine is the coenzyme thiamine diphosphate. Thiamine triphosphate (ThTP) is also found in low amounts in most vertebrate tissues and can phosphorylate certain proteins. Here we show that ThTP exists not only in vertebrates but is present in bacteria, fungi, plants and invertebrates. Unexpectedly, we found that in Escherichia coli as well as in Arabidopsis thaliana, ThTP was synthesized only under particular circumstances such as hypoxia (E. coli) or withering (A. thaliana). In mammalian tissues, ThTP concentrations are regulated by a specific thiamine triphosphatase that we have recently characterized. This enzyme was found only in mammals. In other organisms, ThTP can be hydrolyzed by unspecific phosphohydrolases. The occurrence of ThTP from prokaryotes to mammals suggests that it may have a basic role in cell metabolism or cell signaling. A decreased content may contribute to the symptoms observed during thiamine deficiency.Received 7 March 2003; received after revision 11 April 2003; accepted 14 April 2003     

Thiamine pyrophosphate: An essential cofactor for the α-oxidation in mammals – implications for thiamine deficiencies?

The identification of 2-hydroxyphytanoyl-CoA lyase (2-HPCL), a thiamine pyrophosphate (TPP)-dependent peroxisomal enzyme involved in the α-oxidation of phytanic acid and of 2-hydroxy straight chain fatty acids, pointed towards a role of TPP in these processes. Until then, TPP had not been implicated in mammalian peroxisomal metabolism. The effect of thiamine deficiency on 2-HPCL and α-oxidation has not been studied, nor have possible adverse effects of deficient α-oxidation been considered in the pathogenesis of diseases associated with thiamine shortage, such as thiamine-responsive megaloblastic anemia (TRMA). Experiments with cultured cells and animal models showed that α-oxidation is controlled by the thiamine status of the cell/tissue/organism, and suggested that some pathological consequences of thiamine starvation could be related to impaired α-oxidation. Whereas accumulation of phytanic acid and/or 2-hydroxyfatty acids or their α-oxidation intermediates in TRMA patients given a normal supply of thiamine is unlikely, this may not be true when malnourished. Received 23 December 2005; received after revision 10 April 2006; accepted 28 April 2006     

Active transport of dimethialium inSaccharomyces cerevisiae

Summary Dimethialium, a derivative of thiamine which has a methyl group in place of hydroxyethyl group at the 5-position of the thiazole moiety, was found to be accumulated in nonproliferating cells ofSaccharomyces cerevisiae by the same transport mechanism for thiamine. The results strongly support the supposition that thiamine as well as dimethialium can be transported and accumulated without obligatory phosphorylation in yeast cells, since dimethialium is not phosphorylated by yeast thiamine pyrophosphokinase.We wish to thank the late Dr S. Yurugi, Takeda Research Laboratories, for a generous gift of dimethialium.     

Interaction of basic dyes with the thiamine transport system inSaccharomyces cerevisiae

Summary Basic dyes such as methylene blue and triphenyltetrazolium chloride were found to inhibit thiamine transport inSaccharomyces cerevisiae. Conversely, the reduction of methylene blue and triphenyltetrazolium chloride by yeast cells was inhibited by thiamine. A thiamine transport mutant ofSaccharomyces cerevisiae showed decreased utilization of these dyes. From the results, the possibility that the uptake of basic dyes may proceed via a membrane-bound thiamine-binding protein in the thiamine transport system of the yeast is discussed.This work was supported in part by a grant from the Ministry of Education, Science and Culture of Japan.     

Thiamine-binding activity ofSaccharomyces cerevisiae plasma membrane

Summary The specific binding activity to [¹⁴C]thiamine was found to be located in the plasma membrane ofSaccharomyces cerevisiae. The activity was inhibited by several thiamine analogs and it was hardly detectable in the plasma membrane from a thiamine transport mutant ofSaccharomyces cerevisiae. Some properties of the thiamine-binding activity of yeast plasma membrane are discussed in connection with those of the thiamine transport system.     

Effects of hypochlorite on thiamine and its derivatives

Thiamine and its phosphorylated derivatives reacted with hypochlorite with reaction rates following the order: thiamine greater than thiamine monophosphate greater than thiamine diphosphate from pH 4.0 to 6.5. At least one unknown transient intermediate was formed and at least one non-thiochrome product was fluorescent. Chemiluminescence was also observed.     

Protective effect of vitamins against trichothecene toxicity towards Saccharomyces cerevisiae

Several trichothecene mycotoxins were shown to inhibit the growth of Saccharomyces cerevisiae. This effect was most pronounced with the macrocyclic trichothecenes, especially verrucarin A. Much less growth inhibition was observed with T-2 toxin. Verrucarol, diacetoxyscirpenol, acetyl T-2 toxin, HT-2 toxin, T-2 tetraol and neosolaniol were inactive at a concentration of 75 micrograms of toxin per disc. Incubation of S. cerevisiae with verrucarin A together with vitamins resulted in a decrease in toxicity. Pyridoxine-HCl, Ca-pantothenate, thiamine-HCl and alpha-tocopheryl acetate were amongst the most potent of the vitamins tested which reversed growth inhibition, overcoming the inhibitory potential of the toxins.     

Active transport of [32P]thiamine diphosphate inEscherichia coli

Summary Active uptake of [³²P]thiamine diphosphate byE. coli was analyzed using an improved method of gel filtration chromatography. The radioactive coenzyme was accumulated without dephosphorylation. From this result it was concluded that thiamine kinase is not involved in the membrane transport of thiamine inE. coli.We are indebted to Miss M. Abe for her technical assistance.     

Pharmakologische Untersuchungen bei Thiaminmangel I. Änderungen des Katecholamingehalts im Gewebe

Summary (1) The increased tissue catecholamine level of the thiamine deficient rat was shown in the atrium, the ventricle, the brain cortex and in the spleen but not in the brain stem and the adrenal gland. (2) The increased response to tyramine of the thiamine deficient heart is most probably due to the high catecholamine level caused by thiamine deficiency. (3) The increased catecholamine content in the thiamine deficient rat does not result from the increased pyruvic acid.

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Herrn Professor Dr.F. Th. von Brücke zum 60. Geburtstag im Januar 1968 gewidmet.     

Effect of lipophilic cations on thiamine transport system in isolated rat hepatocytes

The effect of lipophilic cations such as triphenylmethylphosphonium, tetraphenylphosphonium and tetraphenylarsonium in addition to dibenzyldimethylammonium on thiamine transport in isolated rat hepatocytes was studied. Lipophilic cations at the concentration 10 microM almost completely inhibited thiamine uptake. Kinetic studies showed that these compounds were competitive inhibitors with a very high affinity. These results suggest that lipophilic cations in addition to quaternary ammonium compounds also share a common binding site for thiamine in isolated rat hepatocytes.     

Occurrence of thiaminase II inSaccharomyces cerevisiae

Summary It was found that cell-free extracts ofSaccharomyces cerevisiae contain thiaminase II which hydrolyzes thiamine and thiamine analogs. The possible involvement of this enzyme and thiamine-synthesizing enzymes in thiamine production from thiamine antagonists is discussed.30 September 1986Acknowledgments. We thank the late Dr S. Yurugi, Takeda Pharmaceutical Industries, Osaka, for his generous gifts of dimethialium, 2-northiamine, -hydroxyethylthiamine, hydroxymethylpyrimidine, 2-norhydroxymethylpyrimidine and hydroxyethylthiazole. We also thank Prof. H. Nakayama, Yamaguchi Women's College, for his kind supply ofEscherichia coli 70–17 and 26–43.     

Physiological studies on the effects of nutritional imbalance on the central nervous system. II. Effects of thiamine deficiency on oxidative enzymes in the brain of chicken, Gallus domesticus.

The activitiy levels of succinate dehydrogenase, glutamate dehydrogenase and pyruvate dehydrogenase in the fore, mid and hind brain regions of the thiamine deficient chicken, Gallus domesticus were determined. The activity levels of succinate dehydrogenase and glutamate dehydrogenase in all the 3 regions of brain showed augmentation on inducing thiamine deficiency. In contrast the activity levels of pyruvate dehydrogenase decreased in the brain of thiamine deficient animals. It is suggested that these changes in the oxidative enzymes indicate disturbance caused in the operation of the tricarboxylic acid cycle in thiamine deficiency.     

New mitochondrial carriers: an overview

The transport of metabolites, nucleotides and cofactors across the mitochondrial inner membrane is performed by members of mitochondrial carrier family (MCF). These proteins share marked structural features that have made feasible the functional characterization of numerous MCs in the last years. The MCs responsible for transport activities in mitochondria known for decades such as glutamate uptake or ATP-Mg/Pi exchange have recently been identified as well as novel carriers such as those involved in S-adenosylmethionine or thiamine pyrophosphate uptake. Here, after a brief review of the novel data on structural characteristics and import mechanisms of MCF members, we present an exhaustive compilation of human MC sequences, including previously characterized carriers, together with their respective Saccharomyces cerevisiae orthologues, ordered according to the phylogenetic analysis of el Moualij and co-workers [Yeast (1997) 13: 573-581]. We have detected the existence of at least 49 human MC sequences, including those of yet unknown function. An overview of novel MCF members functionally characterized in recent years in mammals and in yeast genomes is presented.     

Physiological studies on the effects of nutritional imbalance on the central nervous system. II. Effects of thiamine deficiency on oxidative enzymes in the brain of chicken,Gallus domesticus

Summary The activity levels of succinate dehydrogenase, glutamate dehydrogenase and pyruvate dehydrogenase in the fore, mid and hind brain regions of the thiamine deficient chicken,Gallus domesticus were determined. The activity levels of scccinate dehydrogenase and glutamate dehydrogenase in all the 3 regions of brain showed augmentation on inducing thiamine deficiency. In contrast the activity levels of pyruvate dehydrogenase decreased in the brain of thiamine deficient animals. It is suggested that these changes in the oxidative enzymes indicate disturbance caused in the operation of the tricarboxylic acid cycle in thiamine deficiency.     

Effects of hypochlorite on thiamine and its derivatives

Summary Thiamine and its phosphorylated derivatives reacted with hypochlorite with reaction rates following the order: thiamine>thiamine monophosphate>thiamine diphosphate from pH4.0 to 6.5. At least one unknown transient intermediate was formed and at least one non-thiochrome product was fluorescent. Chemiluminescence was also observed.We thank the International Foundation for Science, Sweden, and the Thai University Development Council for financial support.     

Enzymhistochemische Untersuchungen an durstaktivierten neurosekretorischen Zellen der Ratte

Summary In hypothalamic neurosecretory nuclei of rats subject to water withdrawl for 1–14 days, a marked increase in the activities of histochemically detected acid phosphatase, thiamine pyrophosphatase and glucose-6-phosphate dehydrogenase is observed. LDH and SDH activities do not change. Two different cell types can be distinguished by means of the acid phosphatase and TPPase reactions.