Skip to main content
SpringerLink
Log in

Xylose metabolism in the anaerobic fungus Piromyces sp. strain E2 follows the bacterial pathway

  • Original Paper
  • Published:
Archives of Microbiology Aims and scope Submit manuscript

Abstract

The anaerobic fungus Piromyces sp. strain E2 metabolizes xylose via xylose isomerase and d-xylulokinase as was shown by enzymatic and molecular analyses. This resembles the situation in bacteria. The clones encoding the two enzymes were obtained from a cDNA library. The xylose isomerase gene sequence is the first gene of this type reported for a fungus. Northern blot analysis revealed a correlation between mRNA and enzyme activity levels on different growth substrates. Furthermore, the molecular mass calculated from the gene sequence was confirmed by gel permeation chromatography of crude extracts followed by activity measurements. Deduced amino acid sequences of both genes were used for phylogenetic analysis. The xylose isomerases can be divided into two distinct clusters. The Piromyces sp. strain E2 enzyme falls into the cluster comprising plant enzymes and enzymes from bacteria with a low G+C content in their DNA. The d-xylulokinase of Piromyces sp. strain E2 clusters with the bacterial d-xylulokinases. The xylose isomerase gene was expressed in the yeast Saccharomyces cerevisiae, resulting in a low activity (25±13 nmol min−1mg protein-1). These two fungal genes may be applicable to metabolic engineering of Saccharomyces cerevisiae for the alcoholic fermentation of hemicellulosic materials.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1.
Fig. 2.
Fig. 3.
Fig. 4.

Similar content being viewed by others

References

  • Akhmanova A, Voncken FGJ, Harhangi H, Hosea KM, Vogels GD Hackstein JHP (1998) Cytosolic enzymes with a mitochondrial ancestry from the anaerobic chytrid Piromyces sp. E2. Mol Microbiol 30:1017–1027

    Article  CAS  PubMed  Google Scholar 

  • Akhmanova A, Voncken FGJ, Hosea KM, Harhangi H, Keltjens JT, Op den Camp HJM, Vogels GD, Hackstein JHP (1999) A hydrogenosome with pyruvate formate-lyase: anaerobic chytrid fungi use an alternative route for pyruvate catabolism. Mol Microbiol 32:1103–1114

    Article  CAS  PubMed  Google Scholar 

  • Amore R, Wilhelm M, Hollenberg CP (1989) The fermentation of xylose—an analysis of the expression of Bacillus and Actinoplanes xylose isomerase genes in yeast. Appl Microbiol Biotechnol 30:351–357

    CAS  Google Scholar 

  • Banerjee S, Archana A, Satyanarayana T (1994) Xylose metabolism in a thermophilic mould Malbranchea pulchella var. sulfurea TMD-8. Curr Microbiol 29:349–352

    CAS  Google Scholar 

  • Borneman WS, Akin DE, Ljungdahl LG (1989) Fermentation products and plant cell wall degrading enzymes produced by monocentric and polycentric anaerobic ruminal fungi. Appl Environ Microbiol 55:1066–1073

    CAS  PubMed  Google Scholar 

  • Brownlee AG (1994) The nucleic acids of anaerobic fungi. In: Mountfort DO, Orpin CG (eds) Anaerobic fungi. Biology, ecology, and function. Marcel Dekker, New York, pp 241–256

  • Bruinenberg PM, de Bot PHM, Van Dijken JP, Scheffers WA (1983) The role of the redox balance in the anaerobic fermentation of xylose by yeasts. Eur J Appl Microbiol Biotechnol 18:287–292

    CAS  Google Scholar 

  • Bruinenberg PM, de Bot PHM, Van Dijken JP, Scheffers WA (1984) NADH-linked aldose reductase: the key to alcoholic fermentation of xylose by yeasts. Appl Microbiol Biotechnol 19:256–269

    CAS  Google Scholar 

  • Chirgwin JM, Przybyla AE, MacDonald RJ, Rutter WJ (1979) Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 18:5294–5299

    CAS  PubMed  Google Scholar 

  • Da Silva ACR, Ferro JA, Reinach FC, Farah CS, Furlan LR, Quaggio RB, Monteiro-Vitorello CB, Van Sluys MA, Almeida Jr NF, Alves LMC, do Amaral AM, Bertolini MC, Camargo LEA, Camarotte G, Cannavan F, Cardozo J, Chambergo F, Ciapina LP, Cicarelli RMB, Coutinho LL, Cursino-Santos JR, El-Dorry H, Faria JB, Ferreira AJS, Ferreira RCC, Ferro MIT, Formighieri EF, Franco MC, Greggio CC, Gruber A, Katsuyama AM, Kishi LT, Leite Jr RP, Lemos EGM, Lemos MVF, Locali EC, Machado MA, Madeira AMBN, Martinez-Rossi NM, Martins EC, Meidanis J, Menck CFM, Miyaki CY, Moon DH, Moreira LM, Novo MTM, OkuraVK, Oliveira MC, Oliveira VR, Pereira Jr HA, Rossi A, Sena JAD, Silva C, de Souza RF, Spinola LAF, Takita MA, Tamura RE, Teixeira EC, Tezza RID, Trindade dos Santos M, Truffi D, Tsai SM, White FF, Setubal JC, Kitajima JP (2002) Comparison of the genomes of two Xanthomonas pathogens with differing host specificities. Nature 417:459–463

    Article  PubMed  Google Scholar 

  • Devereux J, Haeberli P, Smithies O (1984) A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res 12:387–395

    PubMed  Google Scholar 

  • Dische Z, Borenfreund E (1951) A new spectrophotometric method for the detection and determination of keto sugars and trioses. J Biol Chem 192:583–587

    CAS  Google Scholar 

  • Felsenstein J (1989) PHYLIP—Phylogeny Interference Package (Version 3.2). Cladistics 5:164–166

    Google Scholar 

  • Fraenkel DG (1987) Glycolysis, pentose phosphate pathway, and Entner-Doudoroff pathway. In: Neidhardt FC (ed) Escherichia coli and Salmonella typhimurium - cellular and molecular biology. ASM, Washington DC, pp 142–150

  • Fraser CM, Casjens S, Huang WM, Sutton GG, Clayton RA, Lathigra R, White O, Ketchum KA, Dodson R, Hickey EK, Gwinn M, Dougherty B, Tomb J-F, Fleischmann RD, Richardson D, Peterson J, Kerlavage AR, Quackenbush J, Salzberg S, Hanson M, van Vugt R, Palmer N, Adams MD, Gocayne JD, Weidman J, Utterback T, Watthey L, McDonald L, Artiach P, Bowman C, Garland S, Fujii C, Cotton, MD, Horst K, Roberts K, Hatch B, Smith HO, Venter JC (1997) Genomic sequence of a Lyme disease spirochaete, Borrelia burgdorferi. Nature 390:580–586

    Google Scholar 

  • Garcia-Vallvé S, Romeu A, Palau J (2000) Horizontal gene transfer of glycosyl hydrolases of the rumen fungi. Mol Biol Evol 17:352–361

    PubMed  Google Scholar 

  • Gilbert HJ, Hazlewood GP, Laurie JI, Orpin CG, Xue GP (1992) Homologous catalytic domains in a rumen fungal xylanase—Evidence for gene duplication and prokaryotic origin. Mol Microbiol 6:2065–2072

    CAS  PubMed  Google Scholar 

  • Hahn-Hägerdal B, Wahlbom CF, Gárdonyi M, van Zyl WH, Cordero Otero RR, Jönsson LF (2001) Metabolic engineering of Saccharomyces cerevisiae for xylose utilization. Adv Biochem Engin Biotechnol 73:53–84

    Google Scholar 

  • Harhangi HR, Steenbakkers PJM, Akhmanova A, Jetten MSM, van der Drift C, Op den Camp HJM (2002) A highly expressed family 1 β-glucosidase with transglycosylation capacity from the anaerobic fungus Piromyces sp. E2. Biochem Biophys Acta 1574:293–303

    Article  CAS  PubMed  Google Scholar 

  • Harhangi HR, Akhmanova A, Steenbakkers PJM, Jetten MSM, van der Drift C, Op den Camp HJM (2003) Genomic DNA analysis of genes encoding (hemi-)cellulolytic enzymes of the anaerobic fungus Piromyces sp. E2. Gene (in press)

  • Henrick K, Collyer CA, Blow DM (1989) Structures of d-xylose isomerase from Arthrobacter strain B3728 containing the inhibitors xylitol and D-sorbitol at 2.5 Å and 2.3 Å resolution, respectively. J Mol Biol 208:129–157

    CAS  PubMed  Google Scholar 

  • Jeffries TW (1983) Utilization of xylose by bacteria, yeasts, and fungi. Adv Biochem Eng Biotechnol 27:1–32

    CAS  PubMed  Google Scholar 

  • Jeppsson M, Traff K, Johansson B, Hahn-Hägerdal B, Gorwa-Grauslund MF (2003) Effect of enhanced xylose reductase activity on xylose consumption and product distribution in xylose-fermenting recombinant Saccharomyces cerevisiae. FEMS Yeast Res 3:167–175

    Article  CAS  PubMed  Google Scholar 

  • Jin YS, Jeffries T (2003) Changing flux of xylose metabolites by altering expression of xylose reductase and xylitol dehydrogenase in recombinant Saccharomyces cerevisiae. Appl Biochem Biotechnol 106:277–286

    PubMed  Google Scholar 

  • KanekoT, KotaniH, NakamuraY, Sato S, Asamizu E, Miyajima N, Tabata S (1998) Structural analysis of Arabidopsis thaliana chromosome 5. V. Sequence features of the regions of 1,381,565 bp covered by twenty one physically assigned P1 and TAC clones. J DNA Res 5: 131–145

    Google Scholar 

  • Kristo P, Saarelainen R, Fagerstrom R, Aho S, Korhola M (1996) Protein purification, and cloning and characterization of the cDNA and gene for xylose isomerase of barley. Eur J Biochem 237:240–246

    CAS  PubMed  Google Scholar 

  • Kuyper M, Harhangi HR, Stave AK, Winkler AA, Op den Camp HJM, Jetten MSM, de Laat WTAM, den Ridder JJJ, van Dijken JP, Pronk JT (2003) High-level functional expression of a fungal xylose isomerase: the key to efficient alcoholic fermentation of xylose by Saccharomyces cerevisiae? FEMS Yeast Res (in press)

  • Lowe SE, Theodorou MK, Trinci APJ (1987) Growth and fermentation of an anaerobic fungus on various carbon sources and effect of temperature on development. Appl Environ Microbiol 53:1210–1215

    CAS  PubMed  Google Scholar 

  • Marvin-Sikkema FD, Richardson AJ, Stewart CS, Gottschal JC, Prins RA (1990) Influence of hydrogen-consuming bacteria on cellulose degradation by anaerobic fungi. Appl Environ Microbiol 56:3793–3797

    CAS  PubMed  Google Scholar 

  • Marvin-Sikkema FD, Pedro Gomes TM, Grivet JP, Gottschal JC, Prins RA (1993) Characterization of hydrogenosomes and their role in glucose metabolism of Neocallimastix sp. L2. Arch Microbiol 160:388–396

    CAS  PubMed  Google Scholar 

  • Meaden PG, Aduse-Opoku J, Reizer J, Reizer A, Lanceman YA, Martin MF, Mitchell WJ (1994) The xylose isomerase-encoding gene (xylA) of Clostridium thermosaccharolyticum: Cloning, sequencing and phylogeny of XylA enzymes. Gene 141:97–101

    Article  CAS  PubMed  Google Scholar 

  • Rawat U, Phadtare S, Deshpande V, Rao M (1996) A novel xylose isomerase from Neurospora crassa. Biotechnol Lett 18:1267–1270

    CAS  Google Scholar 

  • Richard P, Londesborough J, Putkonen M, Kalkkinen N, Penttila M (2001) Cloning and expression of a fungal L-arabinitol 4-dehydrogenase gene. J Biol Chem 276:40631–40637

    Article  CAS  PubMed  Google Scholar 

  • Rodriguez-Peña JM, Cid VJ, Arroyo J, Nombela C (1998) The YGR194c (XKS1) gene encodes the xylulokinase from the budding yeast Saccharomyces cerevisiae. FEMS Microbiol Lett 162:155–160

    Article  PubMed  Google Scholar 

  • Teunissen MJ, Op den Camp HJM (1993) Anaerobic fungi and their cellulolytic and xylanolytic enzymes. Antonie van Leeuwenhoek 63:63–76

    CAS  PubMed  Google Scholar 

  • Teunissen MJ, Op den Camp HJM, Orpin CG, Huis in 't Veld JHJ, Vogels GD (1991) Comparison of growth characteristics of anaerobic fungi from ruminant and non-ruminant herbivores during growth in a defined medium. J Gen Microbiol 137:1401–1408

    CAS  PubMed  Google Scholar 

  • Trinci APJ, Davies DR, Gull K, Lawrence MI, Nielsen BB, Rickers A, Theodorou MK (1994) Anaerobic fungi in herbivorous animals. Mycol Res 98:129–152

    Google Scholar 

  • Vangrysperre W, Van Damme J, Vandekerckhove J, De Bruyne CK, Cornelis R, Kersters-Hilderson H (1989) Localization of the essential histidine and carboxylate group in xylose isomerases. Biochem J 265:699–705

    Google Scholar 

  • Van Kuyk PA, de Groot MJ, Ruijter GJ, de Vries RP, Visser J (2001) The Aspergillus niger d-xylulose kinase gene is co-expressed with genes encoding arabinan degrading enzymes, and is essential for growth on d-xylose and l-arabinose. Eur J Biochem 268:5414–5423

    Article  CAS  PubMed  Google Scholar 

  • Walfridsson M, Bao X, Anderlund M, Lilius G, Bulow L, Hahn-Hägerdal B (1996) Ethanolic fermentation of xylose with Saccharomyces cerevisiae harboring the Thermus thermophilus xylA gene, which expresses an active xylose (glucose) isomerase. Appl Environ Microbiol 62:4648–4651

    CAS  PubMed  Google Scholar 

  • Wang T, Penttila M, Gao P (1999) Expression of xylose-metabolic key genes of Trichoderma reesei on various carbon sources measured by a series of Northern hybridizations. Wei Sheng Wu Xue Bao 39:503–509

    CAS  PubMed  Google Scholar 

  • Wang T, Penttila M, Gao P, Wang C, Zhong L (1998) Isolation and identification of xylitol dehydrogenase gene from Trichoderma reesei. Chin J Biotechnol 14:179–185

    Article  PubMed  Google Scholar 

  • Williams AG, Orpin CG (1987) Polysaccharide degrading enzymes formed by anaerobic rumen fungi grown on a range of carbohydrate substrates. Can J Microbiol 33:418–426

    CAS  PubMed  Google Scholar 

  • Witteveen CFB, Busink R, van de Vondervoort P, Dijkema C, Swart K, Visser J (1989) l–Arabinose and d-xylose catabolism in Aspergillus niger. J Gen Microbiol 135:2163–2171

    CAS  Google Scholar 

  • Yarlett N, Orpin CG, Munn EA, Yarlett NC, Greenwood CA (1986) Hydrogenosomes in the anaerobic fungus Neocallimastix patriciarum. Biochem J 236:729–739

    CAS  PubMed  Google Scholar 

  • Zhou LQ, Xue GP, Orpin CG, Black GW, Gilbert HJ, Hazlewood GP (1994) Intronless Celb from the anaerobic fungus Neocallimastix patriciarum encodes a modular family A endoglucanase. Biochem. J 297:359–364

    Google Scholar 

Download references

Acknowledgements

We thank Nick van Bakel for excellent technical assistance with the biochemical work and Royal Nedalco B.V. for support. We are grateful to Peter Steenbakkers and Jan Keltjens for critically reading the manuscript and stimulating discussions.

Author information

Author notes

  1. Anna S. Akhmanova

    Present address: Department of Cell Biology and Genetics, Erasmus University, P.O. Box 1738, 3000 , Rotterdam, The Netherlands

Authors and Affiliations

  1. Department of Microbiology, University of Nijmegen, Toernooiveld 1, Faculty of Science, 6525 ED , Nijmegen, The Netherlands

    Harry R. Harhangi, Anna S. Akhmanova, Roul Emmens, Chris van der Drift, Mike S. M. Jetten & Huub J. M. Op den Camp

  2. Royal Nedalco B.V., Postbus 6, 4600 , AA Bergen op Zoom, The Netherlands

    Wim T. A. M. de Laat

  3. Kluyver Laboratory of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 , BC Delft, The Netherlands

    Johannes P. van Dijken & Jack T. Pronk

Authors
  1. Harry R. Harhangi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Anna S. Akhmanova
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Roul Emmens
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Chris van der Drift
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Wim T. A. M. de Laat
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Johannes P. van Dijken
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Mike S. M. Jetten
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Jack T. Pronk
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Huub J. M. Op den Camp
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Mike S. M. Jetten.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Harhangi, H.R., Akhmanova, A.S., Emmens, R. et al. Xylose metabolism in the anaerobic fungus Piromyces sp. strain E2 follows the bacterial pathway. Arch Microbiol 180, 134–141 (2003). https://doi.org/10.1007/s00203-003-0565-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00203-003-0565-0

Keywords

Search

Navigation