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.     

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.     

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.     

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.     

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.     

Effect of growth factor deficiency on nystatin sensitivity inSaccharomyces cerevisiae

Summary The nystatin sensitivity, as well as the levels of total sterols and individual phospholipids ofSaccharomyces cerevisiae are influenced by variations in the supply of growth factors.     

Baker's yeast,an attractant for baiting traps for Chagas' disease vectors

We tested the attraction of volatile compounds, produced by the aerobic growth ofSaccharomyces cerevisiae on saccharose forTriatoma infestans. For these tests, we exploited the behavioural characteristic of these haematophagous insects of dropping when searching for food. In olfactometer assays, yeast cultures activated and attracted bugs as effectively as a mouse. The attraction of the cultures was significantly reduced when the carbon dioxide released was partially eliminated using potassium hydroxide. Yeast cultures were also tested as lures in a novel trap device. A baited device for trapping Chagas' disease vectors using the behavioural peculiarities ofT. infestans and this simple attractant is described.     

Incorporation of photolabile azido fatty acid probes inNeurospora crassa andSaccharomyces cerevisiae

Summary Photolabile azido fatty acids supported the growth of fatty acid auxotrophs, thecel mutant ofNeurospora crassa and the KD115 mutant ofSaccharomyces cerevisiae. These fatty acids were incorporated into the membrane lipids of thecel and KD115 mutants, but were not used by the wild type strain.This work was presented at the 26th International Congress of Pure and Applied Chemistry, Session 1, Tokyo, September 4–10, 1977, abstracts p. 241.Support was provided by the Govt. of India to B.C. and by NCERT, New Delhi India to D.N.C.To whom requests for reprints should be sent.     

Protective effect of vitamins against trichothecene toxicity towardsSaccharomyces cerevisiae

Summary Several trichothecene mycotoxins were shown to inhibit the growth ofSaccharomyces 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 g of toxin per disc. Incubation ofS. cerevisiae with verrucarin A together with vitamins resulted in a decrease in toxicity. Pyridoxine-HCl, Ca-pantothenate, thiamine-HCl and -tocopheryl acetate were amongst the most potent of the vitamins tested which reversed growth inhibition, overcoming the inhibitory potential of the toxins.9 December 1986The authors thank Dr J. Behrend, Makor Company, Israel, for a generous gift of verrucarin A and roridin A.     

Mg2+-ATPase defective mutant ofEscherichia coli and thiamine transport

Summary Mg²⁺-ATPase deficient mutant ofEscherichia coli showed an evident dependency of thiamine uptake on the oxidative metabolism of glucose, whereas the parent strain did not. In both cells, this uptake was completely inhibited by H⁺ conductors.Acknowledgment. We are indebted to Miss M. Abe for her excellent technical assistance.     

Heterozygous effects on cell yield and generation time inSaccharomyces cerevisiae

Summary The data obtained analyzing generation time, cell yield and their variability in different culture media in diploid strains ofSaccharomyces cerevisiae demonstrate the existence of a biochemically determined heterotic effect, that could be of some relevance for the study of yeast population genetics, as well as for the improvement of microbial fermentation processes.     

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     

Localization of wheat germ agglutinin receptor sites on yeast cells by scanning electron microscopy

Summary WGA receptor sites on the surface ofSaccharomyces cerevisiae cells pretreated with an - mannanase were localized by gold granules labelled with WGA. The receptor sites were found on the bud scars, the mother cell-bud junction (chitin) and the bud, but not on the mother cell.Acknowledgment. We thank Prof.C. E. Ballou for supplying theArthrobacter strain GJM-1 and MrsM. Weber for the photographic work.     

Reversal of pyrithiamine-induced growth inhibition of Saccharomyces cerevisiae by oxythiamine

Oxythiamine reversed the growth inhibition of Saccharomyces 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 of S. cerevisiae.     

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.     

Flow microcalorimetric bioassay of polyene antibiotics: Interaction with growingSaccharomyces cerevisiae

Summary Microcalorimetric investigation of the interaction of polyene antibiotics with mid-exponential cells of a growing culture ofSaccharomyces cerevisiae has been used as the basis of a bioassay procedure. The assay is rapid, sensitive and reproducible. The results are compared to classical assays and potency ranking orders.This work was supported by the Science Research Council. (Studentship for B.Z. Chowdhry). We are grateful to the Central Research Fund of London University for funds to purchase the flow nephelometer used to support this study.     

Purification and properties of a heat-resistant exotoxin produced byMacrophomina phaseolina (Tassi) Goid in culture

Summary A partially purified preparation of a water-soluble, heat-resistant, nonspecific exotoxin produced by a strain ofMacrophomina phaseolina, isolated fromPhaseolus mungo L. could reduce Cu⁺⁺ ions and produced a red colour with 2,4-dinitrophenyl hydrazine reagent. It caused inhibition of seed germination, wilting of cut seedlings, stunted growth of young seedlings and loss of permeability of the cell membrane. Seedlings ofP. mungo, grown in presence of the toxin showed a slight increase in the contents of protein and total RNA over control, but a significant increase in the specific activities of F-1, 6-BP aldolase and G-6-P isomerase.     

Consensus phylogeny ofDictyostelium

The evolutionary relationship ofDictyostelium discoideum to the yeasts, fungi, plants, and animals is considered on the basis of physiological, morphological and molecular characteristics. Previous analyses of five proteins indicated thatDictyostelium diverged after the yeasts but before the metazoan radiation. However, analyses of the small ribosomal subunit RNA indicated divergence prior to the yeasts. We have extended the molecular phylogenetic analyses to six more proteins and find consistent evidence for a more recent common ancestor with metazoans than yeast. A consensus phylogeny generated from these new results by both distance matrix and parsimony analyses establishesDictyostelum's place in evolution between the yeastsSaccharomyces cerevisiae andSchizzosaccharomyces pombe and the wormCaenorhabditis elegans.     

Inhibitory effect of halocyamine,an antimicrobial substance from ascidian hemocytes,on the growth of fish viruses and marine bacteria

Summary Halocyamine A, an antimicrobial substance isolated from hemocytes of the solitary ascidianHalocynthia roretzi, inhibited in vitro the growth of fish RNA viruses (infectious hematopoietic necrosis virus and infectious pancreatic necrosis virus). Pretreatment of RNA virus with halocyamine A reduced the infectivity of the virus toward host cells. The growth of marine bacteria,Achromobacter aquamarinus andPseudomonas perfectomarinus, was also inhibited by halocyamine A but that ofAlteromonas putrefaciens andVibrio anguillarum was not. These results suggest that halocyamine may have a role in the defense mechanisms ofH. roretzi against marine viruses and bacteria.