Preferences help
enabled [disable] Abstract
Number of results
2015 | 62 | 4 | 841-850
Article title

The sequence diversity and expression among genes of the folic acid biosynthesis pathway in industrial Saccharomyces strains

Title variants
Languages of publication
Folic acid is an important vitamin in human nutrition and its deficiency in pregnant women's diets results in neural tube defects and other neurological damage to the fetus. Additionally, DNA synthesis, cell division and intestinal absorption are inhibited in case of adults. Since this discovery, governments and health organizations worldwide have made recommendations concerning folic acid supplementation of food for women planning to become pregnant. In many countries this has led to the introduction of fortifications, where synthetic folic acid is added to flour. It is known that Saccharomyces strains (brewing and bakers' yeast) are one of the main producers of folic acid and they can be used as a natural source of this vitamin. Proper selection of the most efficient strains may enhance the folate content in bread, fermented vegetables, dairy products and beer by 100% and may be used in the food industry. The objective of this study was to select the optimal producing yeast strain by determining the differences in nucleotide sequences in the FOL2, FOL3 and DFR1 genes of folic acid biosynthesis pathway. The Multitemperature Single Strand Conformation Polymorphism (MSSCP) method and further nucleotide sequencing for selected strains were applied to indicate SNPs in selected gene fragments. The RT qPCR technique was also applied to examine relative expression of the FOL3 gene. Furthermore, this is the first time ever that industrial yeast strains were analysed regarding genes of the folic acid biosynthesis pathway. It was observed that a correlation exists between the folic acid amount produced by industrial yeast strains and changes in the nucleotide sequence of adequate genes. The most significant changes occur in the DFR1 gene, mostly in the first part, which causes major protein structure modifications in KKP 232, KKP 222 and KKP 277 strains. Our study shows that the large amount of SNP contributes to impairment of the selected enzymes and S. cerevisiae and S. pastorianus produce reduced amounts of the investigated metabolite. The results obtained here yield a list of genetically stable yeast strains which can be implemented as a starter culture in the food industry.
Physical description
  • prof. Waclaw Dabrowski Institute of Agricultural and Food Biotechnology, Department of Microbiology, Culture Collection of Industrial Microorganisms, Warsaw, Poland
  • prof. Waclaw Dabrowski Institute of Agricultural and Food Biotechnology, Department of Microbiology, Culture Collection of Industrial Microorganisms, Warsaw, Poland
  • Amberg DC, Burke DJ, Strathern JN (2006) Yeast DNA isolation: Midiprep. Cold Spring Harbor Protocols (1), pdb.prot4147-pdb.prot4147.
  • Bagley PJ, Selhub J (2000) Analysis of folate form distribution by affinity followed by reversed-phase chromatography with electrical detection. Clin Chem 46: 404-411.
  • Bergström A, Simpson JT, Salinas F, Barré B, Parts L, Zia A, Nguyen Ba AN, Moses AM, Louis EJ, Mustonen V, Warringer J, Durbin R, Liti G (2014) A high-definition view of functional genetic variation from natural yeast genomes. Mol Biol Evol 31: 872-888.
  • Bestwick JP, Huttly WJ, Morris JK, Wald NJ (2014) Prevention of neural tube defects: A cross-sectional study of the uptake of folic acid supplementation in nearly half a million women. PLoS ONE 9: e89354.
  • Duarte FL, Baleiras-Couto M (2012) Evaluation of fermenting grape must yeast dynamics by SSCP profiles. Cienc Tec Vitivinic 27: 95-101.
  • Díaz de la Garza R, Quinlivan EP, Klaus SM, Basset GJ, Gregory JF 3rd, Hanson AD (2004) Folate biofortification in tomatoes by engineering the pteridine branch of folate synthesis. Proc Natl Acad Sci USA 101: 13720-13725.
  • Galeote V, Novo M, Salema-Oom M, Brion C, Valerio E, Goncalves P, Dequin S (2010) FSY1, a horizontally transferred gene in the Saccharomyces cerevisiae EC1118 wine yeast strain encodes a high affinity fructose/H+symporter. Microbiology 156: 3754-3761.
  • Gobbetti M, Cagno RD, De Angelis M (2010) Functional microorganisms for functional food quality. Crit Rev Food Sci Nutr 50: 716-727.
  • Guillaume C, Delobel P, Sablayrolles JM, Blondin B (2007) Molecular basis of fructose utilization by the wine yeast Saccharomyces cerevisiae: a mutated HXT3 allele enhances fructose fermentation. Appl Environ Microbiol. 73: 2432-2439.
  • Gujska E, Majewska K (2005) Effect of baking process on added folic acid and endogenous folates stability in wheat and rye breads. Plant Foods Hum Nutr 60: 37-42.
  • Halsted CH (1980) Intestinal absorption and malabsorption of folates. Annu Rev Med 31: 79-87.
  • Hanson AD, Gregory JF (2011) Folate biosynthesis, turnover, and transport in plants. Annu Rev Plant Biol 62: 105-125.
  • Hayashi K, Yandell DW (1993) How sensitive is PCR-SSCP? Hum Mutat 2: 338-346.
  • Hillenmeyer ME, Fung E, Wildenhain J, Pierce SE, Hoon S, Lee W, Giaever G (2008) The chemical genomic portrait of yeast: uncovering a phenotype for all genes. Science 320: 362-365.
  • Hirsch S, de la Maza P, Barrera G, Gattás V, Petermann M, Bunout D (2002) The Chilean flour folic acid fortification program reduces serum homocysteine levels and masks vitamin B-12 deficiency in elderly people. J Nutr 132: 289-291.
  • Hjortmo SB, Hellström AM, Andlid TA (2008) Production of folates by yeasts in Tanzanian fermented togwa. FEMS Yeast Res 8: 781-787.
  • Hjortmo S, Patring J, Jastebova J, Andlid T (2005) Inherent biodiversity of folate content and composition in yeasts. Trends Food Sci Technol 16: 311-316.
  • Hjortmo S, Patring J, Jastrebova J, Andlid T (2008) Biofortification of folates in white wheat bread by selection of yeast strain and process. Int J Food Microbiol 127: 32-36.
  • Janke T, Schwaiger K, Ege M, Fahn C, von Mutius E, Bauer J, Mayer M (2013) Analysis of the fungal flora in environmental dust samples by PCR-SSCP method. Curr Microbiol 67: 156-169.
  • Jagerstad M, Piironen V, Walker C, Ros G, Carnovale E, Holasova M, Nau H (2005) Increasing natural food folates through bioprocessing and biotechnology. Trends Food Sci Technol 16: 298-306.
  • Jordanova A, Kalaydjieva L, Savov A (1997) SSCP analysis: a blind sensitivity trial. Hum Mutat 10: 65-70.
  • Kaczanowski R, Trzeciak L, Kucharczyk K (2001) Multitemperature single-strand conformation polymorphism. Electrophoresis 22: 3539-3545.
  • Kariluoto S, Aittamaa M, Korhola M, Salovaara H, Vahteristo L, Piironen V (2006) Effects of yeasts and bacteria on the levels of folates in rye sourdoughs. Int J Food Microbiol 106: 137-143.
  • Kariluoto S, Edelmann M, Nyström L, Sontag-Strohm T, Salovaara H, Kivelä R, Piironen V (2014) In situ enrichment of folate by microorganisms in beta-glucan rich oat and barley matrices. Int J Food Microbiol 176: 38-48.
  • Kerr JR, Curran MD (1996) Applications of polymerase chain reaction-single stranded conformational polymorphism to microbiology. Clin Mol Pathol 49: M315-M320.
  • LeBlanc JG, Laino JE, del Valle MJ, Vannini V, van Sinderen D, Taranto MP, de Valdez GF, de Giori GS, Sesma F (2001) B-Group vitamin production by lactic acid bacteria - current knowledge and potential applications. J Appl Microbiol 111: 1297-1309.
  • LeBlanc JG, Milani C, de Giori GS, Sesma F, van Sinderen D, Ventura M (2013) Bacteria as vitamin suppliers to their host: a gut microbiota perspective. Curr Opin Biotechnol 24: 160-168.
  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using Real-Time quantitative PCR and the 2−ΔΔCt method. Methods 25: 402-408.
  • Lonn E, Yusuf S, Arnold MJ, Sheridan P, Pogue J, Micks M, McQueen MJ, Probstfield J, Fodor G, Held C, Genest J Jr, Heart Outcomes Prevention Evaluation (HOPE) 2 Investigators (2006) Homocysteine lowering with folic acid and B vitamins in vascular disease. N Engl J Med 354: 1567-1577.
  • Mancini R, Saracino F, Buscemi G, Fischer M, Schramek N, Bracher A, Bacher A, Gütlich M, Carbone ML (1999) Complementation of the fol2 deletion in Saccharomyces cerevisiae by human and Escherichia coli genes encoding GTP Cyclohydrolase I. Biochem Biophys Res Commun 255: 521-527.
  • Martin DW (1983) Water-soluble vitamins. In Harper's review of biochemistry, Martin DW, Mayes PA, Rodwell VW eds, pp 97-113, Lange, Los Atlos, CA.
  • McKillop DJ, Pentieva K, Daly D, McPartlin JM, Hughes J, Strain JJ, Scott JM, McNulty H (2002) The effect of different cooking methods on folate retention in various foods that are amongst the major contributors to folate intake in the UK diet. Br J Nutr 88: 681-688.
  • McLone DG (2003) The etiology of neural tube defects: the role of folic acid. Childs Nerv Syst 19: 537-539.
  • McNulty H, Pentieva K (2004) Folate bioavailability. Proc Nutr Soc 63: 529-536.
  • Misiewicz A (2013) Genetic basis of variability of Saccharomyces industrial brewer's yeast strains. ISBN 978-83-933341-4-8, (in Polish, abstract in English).
  • Misiewicz A, Wróblewska-Kabba S (2014) Adaptative evolution of yeasts genomes from Saccharomyces cerevisiae sensu stricto group. Postep Mikrobiol 53: 211-222, (in Polish, abstract in English).
  • Nardese V, Gütlich M, Brambilla A, Carbone ML (1996) Disruption of the GTP-cyclohydrolase I gene in Saccharomyces cerevisiae. Biochem Biophys Res Commun 218: 273-279.
  • Owens JE, Clifford AJ, Bamforth CW (2007) Folate in beer. J I Brewing 113: 243-248.
  • Patring JD, Hjortmo SB, Jastrebova JA, Svensson UK, Andlid TA, Jägerstad IM (2006) Characterization and quantification of folates produced by yeast strains isolated from kefir granules. Eur Food Res Technol 223: 633-637.
  • Patring JD, Jastrebova JA, Hjortmo SB, Andlid TA, Jägerstad IM (2005) Development of a simplified method for the determination of folates in baker's yeast by HPLC with ultraviolet and fluorescence detection. J Agric Food Chem 53: 2406-2411.
  • Pfeiffer CM, Rogers LM, Bailey LB, Gregory JF (1997) Absorption of folate from fortified cereal-grain products and supplemental folate consumed with or without food, determined by using a dual label stable-isotope protocol. Am J Clin Nutr 66: 1388-1397.
  • Rush D (2000) Nutrition and maternal mortality in the developing world. The Am J Clin Nutr 72: 212s-240s.
  • Schacherer J, Ruderfer DM, Gresham D, Dolinski K, Botstein D, Kruglyak L (2007) Genome-wide analysis of nucleotide-level variation in commonly used Saccharomyces cerevisiae strains. PLoS One 2: e322.
  • Seyoum E, Selhub J (1998) Properties of food folates determined by stability and susceptibility to intestinal pteroylpolyglutamate hydrolase action. J Nutr 128: 1956-1960.
  • Storozhenko S, De Brouwer V, Volckaert M, Navarrete O, Blancquaert D, Zhang GF, Lambert W, Van Der Straeten D (2007) Folate fortification of rice by metabolic engineering. Nat Biotechnol 25: 1277-1279.
  • Stover P (2009) Folate biochemical pathways and their regulation. Folate in health and disease. Second Edn, pp 49-74.
  • Strandler HS, Patring J, Jägerstad M, Jastrebova J (2015) Challenges in the determination of unsubstituted food folates: Impact of stabilities and conversions on analytical results. J Agric Food Chem 63: 2367-2377.
  • Sunnucks P, Wilson AC, Beheregaray LB, Zenger K, French J, Taylor AC (2000) SSCP is not so difficult: the application and utility of single-stranded conformation polymorphism in evolutionary biology and molecular ecology. Mol Ecol 9: 1699-1710.
  • Szewczyk B, Barski P, Sihler W, Rabalski L, Skrzecz I, Hoyos-Carvajal L, de Souza ML (2008) Detection and identification of baculovirus pesticides by multitemperature single-strand conformational polymorphism. J Environ Sci Health B 43: 539-545.
  • Tomczyk-Żak K, Kaczanowski S, Górecka M, Zielenkiewicz U (2012) Novel application of the MSSCP method in biodiversity studies. J Basic Microbiol 52: 104-109.
  • Walkey CJ, Kitts DD, Liu Y, van Vuuren HJJ (2015) Bioengineering yeast to enhance folate levels in wine. Process Biochem 50: 205-210.
  • Walker MJ, Rapley R (2005) Medical Biomethods Handbook, pp 73-75. Humana Press.
  • Wills L (1931) Treatment of pernicious anaemia of pregnancy and tropical anaemia. Br Med J 1(3676): 1059.
  • Winzeler EA, Castillo-Davis CI, Oshiro G, Liang D, Richards DR, Zhou Y, Hartl DL (2003) Genetic diversity in yeast assessed with whole-genome oligonucleotide arrays. Genetics 163: 79-89.
Document Type
Publication order reference
YADDA identifier
JavaScript is turned off in your web browser. Turn it on to take full advantage of this site, then refresh the page.