Abstract
Vitamin B1 (thiamine) is an essential cofactor for several key enzymes of carbohydrate metabolism. Mammals have to salvage this crucial nutrient from their diet to complement their deficiency of de novo synthesis. In contrast, bacteria, fungi, plants and, as reported here, Plasmodium falciparum, possess a vitamin B1 biosynthesis pathway. The plasmodial pathway identified consists of the three vitamin B1 biosynthetic enzymes 5-(2-hydroxy-ethyl)-4-methylthiazole (THZ) kinase (ThiM), 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP)/HMP-P kinase (ThiD) and thiamine phosphate synthase (ThiE). Recombinant PfThiM and PfThiD proteins were biochemically characterised, revealing Kmapp values of 68 μM for THZ and 12 μM for HMP. Furthermore, the ability of PfThiE for generating vitamin B1 was analysed by a complementation assay with thiE-negative E. coli mutants. All three enzymes are expressed throughout the developmental blood stages, as shown by Northern blotting, which indicates the presence of the vitamin B1 biosynthesis enzymes. However, cultivation of the parasite in minimal medium showed a dependency on the provision of HMP or thiamine. These results demonstrate that the human malaria parasite P. falciparum possesses active vitamin B1 biosynthesis, which depends on external provision of thiamine precursors.
References
Blattner, F.R., Plunkett, G. III, Bloch, C.A., Perna, N.T., Burland, V., Riley, M., Collado-Vides, J., Glasner, J.D., Rode, C.K., Mayhew, G.F., et al. (1997). The complete genome sequence of Escherichia coli K-12. Science277, 1453–1474.10.1126/science.277.5331.1453Search in Google Scholar
Begley, T.P., Downs, D.M., Ealick, S.E., McLafferty, F.W., Van Loon, A.P., Taylor, S., Campobasso, N., Chiu, H.J., Kinsland, C., Reddick, J.J., and Xi, J. (1999). Thiamin biosynthesis in prokaryotes. Arch. Microbiol.171, 293–300.10.1007/s002030050713Search in Google Scholar
Bozdech, Z. and Ginsburg, H. (2005). Data mining of the transcriptome of Plasmodium falciparum: the pentose phosphate pathway and ancillary processes. Malar. J.4, 17.10.1186/1475-2875-4-17Search in Google Scholar
Bozdech, Z., Llinas, M., Pulliam, B.L., Wong, E.D., Zhu, J., and DeRisi, J.L. (2003). The transcriptome of the intraerythrocytic developmental cycle of Plasmodium falciparum. PLoS Biol.1, E5.10.1371/journal.pbio.0000005Search in Google Scholar
Bradford, M.M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem.72, 248–254.10.1016/0003-2697(76)90527-3Search in Google Scholar
Campobasso, N., Mathews, I.I., Begley, T.P., and Ealick, S.E. (2000). Crystal structure of 4-methyl-5-β-hydroxyethylthiazole kinase from Bacillus subtilis at 1.5 Å resolution. Biochemistry39, 7868–7877.10.1021/bi0000061Search in Google Scholar
Cassera, M.B., Gozzo, F.C., D'Alexandri, F.L., Merino, E.F., del Portillo, H.A., Peres, V.J., Almeida, I.C., Eberlin, M.N., Wunderlich, G., Wiesner, J., et al. (2004). The methylerythritol phosphate pathway is functionally active in all intraerythrocytic stages of Plasmodium falciparum. J. Biol. Chem.279, 51749–51759.10.1074/jbc.M408360200Search in Google Scholar
Cheng, G., Bennett, E.M., Begley, T.P., and Ealick, S.E. (2002). Crystal structure of 4-amino-5-hydroxymethyl-2-methylpyrimidine phosphate kinase from Salmonella typhimurium at 2.3 Å resolution. Structure (Camb.)10, 225–235.10.1016/S0969-2126(02)00708-6Search in Google Scholar
Chiu, H.J., Reddick, J.J., Begley, T.P., and Ealick, S.E. (1999). Crystal structure of thiamin phosphate synthase from Bacillus subtilis at 1.25 Å resolution. Biochemistry38, 6460–6470.10.1021/bi982903zSearch in Google Scholar PubMed
Das Gupta, R., Krause-Ihle, T., Bergmann, B., Müller, I.B., Khomutov, A.R., Müller, S., Walter, R.D., and Lüersen, K. (2005). 3-Aminooxy-1-aminopropane and derivatives have an antiproliferative effect on cultured Plasmodium falciparum by decreasing intracellular polyamine concentrations. Antimicrob. Agents Chemother.49, 2857–2864.10.1128/AAC.49.7.2857-2864.2005Search in Google Scholar PubMed PubMed Central
Drautz, H., Messerer, W., Zahner, H., Breiding-Mack, S., and Zeeck, A. (1987). Metabolic products of microorganisms. 239. Bacimethrin isolated from Streptomyces albus: identification, derivatives, synthesis and biological properties. J. Antibiot. (Tokyo)40, 1431–1499.Search in Google Scholar
Friedrich, W. (1988). Vitamins (Berlin, New York: Walter de Gruyter).Search in Google Scholar
Gardner, M.J., Hall, N., Fung, E., White, O., Berriman, M., Hyman, R.W., Carlton, J.M., Pain, A., Nelson, K.E., Bowman, S., et al. (2002). Genome sequence of the human malaria parasite Plasmodium falciparum. Nature419, 498–511.10.1038/nature01097Search in Google Scholar
Hohmann, S. and Meacock, P.A. (1998). Thiamin metabolism and thiamin diphosphate-dependent enzymes in the yeast Saccharomyces cerevisiae: genetic regulation. Biochim. Biophys. Acta1385, 201–219.10.1016/S0167-4838(98)00069-7Search in Google Scholar
Imamura, N. and Nakayama, H. (1982). ThiK and ThiL loci of Escherichia coli. J. Bacteriol.151, 708–717.10.1128/jb.151.2.708-717.1982Search in Google Scholar
Ito, S., Fushinobu, S., Yoshioka, I., Koga, S., Matsuzawa, H., and Wakagi, T. (2001). Structural basis for the ADP-specificity of a novel glucokinase from a hyperthermophilic archaeon. Structure (Camb.)9, 205–214.10.1016/S0969-2126(01)00577-9Search in Google Scholar
Kim, Y.S., Nosaka, K., Downs, D.M., Kwak, J.M., Park, D., Chung, I.K., and Nam, H.G. (1998). A Brassica cDNA clone encoding a bifunctional hydroxyl-methylpyrimidine kinase/thiamin-phosphate pyrophosphorylase involved in thiamin biosynthesis. Plant. Mol. Biol.37, 955–966.Search in Google Scholar
Kyes, S., Pinches, R., and Newbold, C. (2000). A simple RNA analysis method shows var and rif multigene family expression patterns in Plasmodium falciparum. Mol. Biochem. Parasitol.105, 311–315.10.1016/S0166-6851(99)00193-0Search in Google Scholar
Lambros, C. and Vanderberg, J.P. (1979). Synchronization of Plasmodium falciparum erythrocytic stages in culture. J. Parasitol.65, 418–42010.2307/3280287Search in Google Scholar
Lawhorn, B.G., Gerdes, S.Y., and Begley, T.P. (2004). A genetic screen for the identification of thiamin metabolic genes. J. Biol. Chem.279, 43555–43559.10.1074/jbc.M404284200Search in Google Scholar PubMed
Le Roch, K.G., Zhou, Y., Blair, P.L., Grainger, M., Moch, J.K., Haynes, J.D., De La Vega, P., Holder, A.A., Batalov, S., Carucci, D.J., and Winzeler, E.A. (2003). Discovery of gene function by expression profiling of the malaria parasite life cycle. Science301, 1503–1508.10.1126/science.1087025Search in Google Scholar PubMed
Li, M.H., Kwok, F., Chang, W.R., Lau, C.K., Zhang, J.P., Lo, S.C., Jiang T., and Liang, D.C. (2002). Crystal structure of brain pyridoxal kinase, a novel member of the ribokinase superfamily. J. Biol. Chem.277, 46385–46390.10.1074/jbc.M208600200Search in Google Scholar PubMed
Llorente, B., Fairhead, C., and Dujon, B. (1999). Genetic redundancy and gene fusion in the genome of the baker's yeast Saccharomyces cerevisiae: functional characterization of a three-member gene family involved in the thiamine biosynthetic pathway. Mol. Microbiol.32, 1140–1152.10.1046/j.1365-2958.1999.01412.xSearch in Google Scholar PubMed
Mathews, I.I., Erion, M.D., and Ealick, S.E. (1998). Structure of human adenosine kinase at 1.5 Å resolution. Biochemistry37, 15607–15620.Search in Google Scholar
Mizote, T. and Nakayama, H. (1989). Purification and properties of hydroxymethyl-pyrimidine kinase from Escherichia coli. Biochim. Biophys. Acta991, 109–113.10.1016/0304-4165(89)90035-4Search in Google Scholar
Mizote, T., Tsuda, M., Nakazawa, T., and Nakayama, H. (1996). The thiJ locus and its relation to phosphorylation of hydroxymethylpyrimidine in Escherichia coli. Microbiology142, 2969–2974.10.1099/13500872-142-10-2969Search in Google Scholar
Mizote, T., Tsuda, M., Smith, D.D., Nakayama, H., and Nakazawa, T. (1999). Cloning and characterization of the thiD/J gene of Escherichia coli encoding a thiamin-synthesizing bifunctional enzyme, hydroxymethylpyrimidine kinase/phosphomethylpyrimidine kinase. Microbiology145, 495–501.10.1099/13500872-145-2-495Search in Google Scholar
Morett, E., Korbel, J.O., Rajan, E., Saab-Rincon, G., Olvera, L., Olvera, M., Schmidt, S., Snel, B., and Bork, P. (2003). Systematic discovery of analogous enzymes in thiamin biosynthesis. Nat. Biotechnol.21, 790–795.10.1038/nbt834Search in Google Scholar
Newell, P.C. and Tucker, R.G. (1968a). Biosynthesis of the pyrimidine moiety of thiamine. A new route of pyrimidine biosynthesis involving purine intermediates. Biochem. J.106, 279–287.10.1042/bj1060279Search in Google Scholar
Newell, P.C. and Tucker, R.G. (1968b). Precursors of the pyrimidine moiety of thiamine. Biochem. J.106, 271–277.10.1042/bj1060271Search in Google Scholar
Nosaka, K., Nishimura, H., Kawasaki, Y., Tsujihara, T., and Iwashima, A. (1994). Isolation and characterization of the THI6 gene encoding a bifunctional thiamin-phosphate pyrophosphorylase/hydroxyethylthiazole kinase from Saccharomyces cerevisiae. J. Biol. Chem.269, 30510–30516.10.1016/S0021-9258(18)43843-4Search in Google Scholar
Park, J.H., Burns, K., Kinsland, C., and Begley, T.P. (2004). Characterization of two kinases involved in thiamine pyrophosphate and pyridoxal phosphate biosynthesis in Bacillus subtilis: 4-amino-5-hydroxymethyl-2methylpyrimidine kinase and pyridoxal kinase. J. Bacteriol.186, 1571–1573.10.1128/JB.186.5.1571-1573.2004Search in Google Scholar PubMed PubMed Central
Peapus, D.H., Chiu, H.J., Campobasso, N., Reddick, J.J., Begley, T.P., and Ealick, S.E. (2001). Structural characterization of the enzyme-substrate, enzyme-intermediate, and enzyme-product complexes of thiamin phosphate synthase. Biochemistry40, 10103–10114.10.1021/bi0104726Search in Google Scholar PubMed
Pohl, M., Sprenger, G.A., and Müller, M. (2004). A new perspective on thiamine catalysis. Curr. Opin. Biotechnol.15, 335–342.10.1016/j.copbio.2004.06.002Search in Google Scholar PubMed
Reddick, J.J., Kinsland, C., Nicewonger, R., Christian, T., Downs, D.M., Winkler, M.E., and Begley, T.P. (1998). Overexpression, purification and characterization of two pyrimidine kinases involved in the biosynthesis of thiamine: 4-amino-5-hydroxymethyl-2-methylpyrimidine kinase and 4-amino-5-hydroxymethyl-2-methy pyrimidine phosphate kinase. Tetrahedron54, 15983–15991.10.1016/S0040-4020(98)01006-0Search in Google Scholar
Reddick, J.J., Saha, S., Lee, J., Melnick, J.S., Perkins, J., Begley, T.P. (2001a). The mechanism of action of bacimethrin, a naturally occurring thiamin antimetabolite. Bioorg. Med. Chem. Lett.11, 2245–2248.10.1016/S0960-894X(01)00373-0Search in Google Scholar
Reddick, J.J., Nicewonger, R., and Begley, T.P. (2001b). Mechanistic studies on thiamin phosphate synthase: evidence for a dissociative mechanism. Biochemistry40, 10095–10102.10.1021/bi010267qSearch in Google Scholar
Rodionov, D.A., Vitreschak, A.G., Mironov, A.A., and Gelfand, M.S. (2002). Comparative genomics of thiamin biosynthesis in procaryotes. New genes and regulatory mechanisms. J. Biol. Chem.277, 48949–48959.10.1074/jbc.M208965200Search in Google Scholar
Sambrook, J., Fritsch, E.F., and Maniatis, T. (1989). Molecular Cloning: A Laboratory Manual, 2nd ed. (Cold Spring Harbor, NY, USA: Cold Spring Harbor Laboratory Press).Search in Google Scholar
Schopfer, W.H. (1949). Plants and Vitamins (Waltham, MA, USA: Chronica Botanica Company).Search in Google Scholar
Scott, T.C. and Phillips, M.A. (1997). Characterization of Trypanosoma brucei pyridoxal kinase: purification, gene isolation and expression in Escherichia coli. Mol. Biochem. Parasitol.88, 1–11.10.1016/S0166-6851(97)00077-7Search in Google Scholar
Sherman, I.W. (1979). Biochemistry of Plasmodium (malarial parasites). Microbiol. Rev.43, 453–495.10.1128/mr.43.4.453-495.1979Search in Google Scholar
Sigrell, J.A., Cameron, A.D., Jones, T.A., and Mowbray, S.L. (1998). Structure of Escherichia coli ribokinase in complex with ribose and dinucleotide determined to 1.8 Å resolution: insights into a new family of kinase structures. Structure (Camb.)6, 183–193.Search in Google Scholar
Stolz, J. and Vielreicher, M. (2003). Tpn1p, the plasma membrane vitamin B6 transporter of Saccharomyces cerevisiae. J. Biol. Chem.278, 18990–18996.10.1074/jbc.M300949200Search in Google Scholar
Taylor, S.V., Kelleher, N.L., Kinsland, C., Chiu, H.J., Costello, C.A., Backstrom, A.D., McLafferty, F.W., and Begley, T.P. (1998). Thiamin biosynthesis in Escherichia coli. Identification of this thiocarboxylate as the immediate sulfur donor in the thiazole formation. J. Biol. Chem.273, 16555–16560.10.1074/jbc.273.26.16555Search in Google Scholar
Tazuya, K., Adachi, Y., Masuda, K., Yamada, K., and Kumaoka, H. (1995). Origin of the nitrogen atom of pyridoxine in Saccharomyces cerevisiae. Biochim. Biophys. Acta1244, 113–116.10.1016/0304-4165(94)00205-CSearch in Google Scholar
Toms, A.V., Haas, A.L., Park, J.H., Begley, T.P., and Ealick, S.E. (2005). Structural characterization of the regulatory proteins TenA and TenI from Bacillus subtilis and identification of TenA as a thiaminase II. Biochemistry44, 2319–1329.10.1021/bi0478648Search in Google Scholar PubMed
Trager, W. and Jensen, J.B. (1976). Human malaria parasites in continuous culture. Science193, 673–675.10.1126/science.781840Search in Google Scholar PubMed
Umlas, J. and Fallon, J.N. (1971). New thick-film technique for malaria diagnosis. Use of saponin stromatolytic solution for lysis. Am. J. Trop. Med. Hyg.20, 527–529.10.4269/ajtmh.1971.20.527Search in Google Scholar PubMed
Wightman, R. and Meacock, P.A. (2003). The THI5 gene family of Saccharomyces cerevisiae: distribution of homologues among the hemiascomycetes and functional redundancy in the aerobic biosynthesis of thiamin from pyridoxine. Microbiology149, 1447–1460.10.1099/mic.0.26194-0Search in Google Scholar PubMed
Wrenger, C., Lüersen, K., Krause, T., Müller, S., and Walter, R.D. (2001). The Plasmodium falciparum bifunctional ornithine decarboxylase, S-adenosyl-l-methionine decarboxylase, enables a well balanced polyamine synthesis without domain-domain interaction. J. Biol. Chem.276, 29651–29656.10.1074/jbc.M100578200Search in Google Scholar PubMed
Wrenger, C., Eschbach, M.L., Müller, I.B., Warnecke, D., and Walter, R.D. (2005). Analysis of the vitamin B6 biosynthesis pathway in the human malaria parasite Plasmodium falciparum. J. Biol. Chem.280, 5242–5248.10.1074/jbc.M412475200Search in Google Scholar PubMed
Zeidler, J., Ullah, N., Gupta, R.N., Pauloski, R.M., Sayer, B.G., and Spenser, I.D. (2002). 2′-Hydroxypyridoxol, a biosynthetic precursor of vitamins B6 and B1 in yeast. J. Am. Chem. Soc.124, 4542–4543.10.1021/ja012708zSearch in Google Scholar PubMed
Zeidler, J., Sayer, B.G., and Spenser, I.D. (2003). Biosynthesis of vitamin B1 in yeast. Derivation of the pyrimidine unit from pyridoxine and histidine. Intermediacy of urocanic acid. J. Am. Chem. Soc.125, 13094–13105.Search in Google Scholar
Zhang, Y., Taylor, S.V., Chiu, H.J., and Begley, T.P. (1997). Characterization of the Bacillus subtilis thiC operon involved in thiamine biosynthesis. J. Bacteriol.179, 3030–3035.10.1128/jb.179.9.3030-3035.1997Search in Google Scholar PubMed PubMed Central
Zilles, J.L., Croal, L.R., and Downs, D.M. (2000). Action of the thiamine antagonist bacimethrin on thiamine biosynthesis. J. Bacteriol.182, 5606–5610.10.1128/JB.182.19.5606-5610.2000Search in Google Scholar PubMed PubMed Central
Zurlinden, A. and Schweingruber, M.E. (1994). Cloning, nucleotide sequence, and regulation of Schizosaccharomyces pombethi4, a thiamine biosynthetic gene. J. Bacteriol.176, 6631–6635.10.1128/jb.176.21.6631-6635.1994Search in Google Scholar PubMed PubMed Central
©2006 by Walter de Gruyter Berlin New York