Skip to main content
SpringerLink
Log in

Sugar repression in the methylotrophic yeastHansenula polymorpha studied by using hexokinase-negative, glucokinase-negative and double kinase-negative mutants

  • Papers
  • Published:
Folia Microbiologica Aims and scope Submit manuscript

Abstract

Two glucose-phosphorylating enzymes, a hexokinase phosphorylating both glucose and fructose, and a glucose-specific glucokinase were electrophoretically separated in the methylotrophic yeastHansenula polymorpha. Hexokinase-negative mutants were isolated inH. polymorpha by using mutagenesis, selection and genetic crosses. Regulation of synthesis of the sugar-repressed alcohol oxidase, catalase and maltase was studied in different hexose kinase mutants. In the wild type and in mutants possessing either hexokinase or glucokinase, glucose repressed the synthesis of maltase, alcohol oxidase and catalase. Glucose repression of alcohol oxidase and catalase was abolished in mutants lacking both glucose-phosphorylating enzymes (i.e. in double kinase-negative mutants). Thus, glucose repression inH. polymorpha cells requires a glucose-phosphorylating enzyme, either hexokinase or glucokinase. The presence of fructose-phosphorylating hexokinase in the cell was specifically needed for fructose repression of alcohol oxidase, catalase and maltase. Hence, glucose or fructose has to be phosphorylated in order to cause repression of the synthesis of these enzymes inH. polymorpha suggesting that sugar repression in this yeast therefore relies on the catalytic activity of hexose kinases.

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

Similar content being viewed by others

References

  • Alamäe T., Simisker J.: Isolation and preliminary characterization ofPichia pinus mutants insensitive to glucose repression.Yeast 10, 1459–1466 (1994).

    Article  PubMed  Google Scholar 

  • Alamäe T., Liiv L.: Glucose repression of maltase and methanol-oxidizing enzymes in the methylotrophic yeastHansenula polymorpha: isolation and study of regulatory mutants.Folia Microbiol. 43, 443–452 (1998).

    Article  Google Scholar 

  • Angell S., Schwarz E., Bibb M.J.: The glucose kinase gene ofStreptomyces coelicolor A3(2): its nucleotide sequence, transcriptional analysis and role in glucose repression.Mol. Microbiol. 52, 2833–2844 (1992).

    Article  Google Scholar 

  • Barnett J.A.: The catabolism of acyclic polyols by yeasts.J. Gen. Microbiol. 52, 131–159 (1968).

    CAS  Google Scholar 

  • van Dijk R., Faber K.N., Kiel J.A.K.W., Veenhuis M., van der Klei I.: The methylotrophic yeastHansenula polymorpha: a versatile cell factory.Enzyme Microb. Technol. 26, 793–800 (2000).

    Article  PubMed  Google Scholar 

  • Eggeling L., Sahm H.: Derepression and partial insensitivity to carbon catabolite repression of the methanol dissimilating enzymes inHansenula polymorpha.Eur. J. Appl. Microbiol. Biotechnol. 5, 197–202 (1978).

    Article  CAS  Google Scholar 

  • Entian K.-D.: Genetic and biochemical evidence for hexokinase PII as a key enzyme involved, in catabolite repression in yeast.Mol. Gen. Genet. 178, 633–637 (1980).

    Article  PubMed  CAS  Google Scholar 

  • Entian K.-D.: Sugar phosphorylation, in yeast, pp. 67–79 in F.K. Zimmermann, K.-D. Entian (Eds):Yeast Sugar Metabolism. Lancaster, Basel 1997.

    Google Scholar 

  • Entian K.-D., Fröhlich K.-U.:Saccharomyces cerevisiae mutants provide evidence of hexokinase PII as a bifunctional enzyme with catalytic and regulatory domains for triggering carbon catabolite repression.J. Bacteriol. 158, 29–35 (1984).

    PubMed  CAS  Google Scholar 

  • Espinel A.E., Gomez-Toribio V., Peinado J.: The inactivation of hexokinase activity does not prevent glucose repression inCandida utilis.FEMS Microbiol. Lett. 135, 327–332 (1995).

    Article  Google Scholar 

  • Gancedo J.M.: Yeast carbon catabolite repression.Microbiol. Mol. Biol. Rev. 62, 334–361 (1998).

    PubMed  CAS  Google Scholar 

  • Gancedo J.M., Clifton D., Fraenkel D.G.: Yeast hexokinase mutants.J. Biol. Chem. 252, 4443–4444 (1977).

    PubMed  CAS  Google Scholar 

  • Gellissen G., Hollenberg C.P.: Application of yeasts in gene expression studies: a comparison ofSaccharomyces cerevisiae, Hansenula polymorpha andKlyuveromyces lactis.Gene 190, 87–97 (1997).

    Article  PubMed  CAS  Google Scholar 

  • Gleeson M.A., Waites M.J., Sudbery P.E.: Development of techniques for genetic analysis in the methylotrophic yeastHansenula polymorpha, pp. 228–235 in R.L. Crawford, R.S. Hanson (Eds).Microbial Growth on C1 Compounds, American Society for Microbiology, Washington (DC) 1984.

    Google Scholar 

  • Goffrini P., Ficarelli A., Ferrero I.: Hexokinase activity is affected in mutants ofKluyveromyces lactis resistant to glucose repression.Microbiology 141, 441–447 (1995).

    CAS  Google Scholar 

  • Heredia M.F., Heredia C.F.:Saccharomyces cerevisiae acquires resistance to 2-deoxyglucose at a very high frequency.J. Bacteriol. 170, 2870–2872 (1988).

    PubMed  CAS  Google Scholar 

  • Herrero P., Galindez J., Ruiz N., Martinez-Campa C., Moreno F.: Transcriptional regulation of theSaccharomyces cerevisiae HXK1, HXK2 andGLK1 genes.Yeast 11, 137–144 (1995).

    Article  PubMed  CAS  Google Scholar 

  • Herrero P., Martinez-Campa C., Moreno F.: The hexokinase 2 protein participates in regulatory DNA-protein complexes necessary for glucose repression of theSUC2 gene inSaccharomyces cerevisiae.FEBS Lett. 434, 71–76 (1998).

    Article  PubMed  CAS  Google Scholar 

  • Hirai M., Ohtani E., Tanaka A., Fukui S.: Glucose-phosphorylating enzymes ofCandida yeasts and their regulationin vivo.Biochim. Biophys. Acta 480, 357–366 (1977).

    PubMed  CAS  Google Scholar 

  • Klein C.J.L., Olsson L., Nielsen J.: Glucose control inSaccharomyces cerevisiae: the role ofMIG1 in metabolic functions.Microbiology 144, 13–24 (1998).

    PubMed  CAS  Google Scholar 

  • Lobo Z., Maitra P.K.: Resistance to 2-deoxyglucose in yeast: a direct selection of mutants lacking glucose-phosphorylating enzymes.Mol. Gen. Genet. 157, 297–300 (1977).

    Article  PubMed  CAS  Google Scholar 

  • Ma H., Botstein D.: Effects of null mutations in the hexokinase genes ofSaccharomyces cerevisiae on catabolite repression.Mol. Cell. Biol. 6, 4046–4052 (1986).

    PubMed  CAS  Google Scholar 

  • Ma H., Bloom L.M., Walsh C.T., Botstein D.: The residual enzymatic phosphorylation activity of hexokinase II mutants is correlated with glucose repression inSaccharomyces cerevisiae.Mol. Cell. Biol. 9, 5643–5649 (1989).

    PubMed  CAS  Google Scholar 

  • Maitra P.K.: A glucokinase fromSaccharomyces cerevisiae.J. Biol. Chem. 245, 2423–2431 (1970).

    PubMed  CAS  Google Scholar 

  • Mazon M., Gancedo J.M., Gancedo C.: Hexose kinases fromRhodotorula glutinis.Arch. Biochem. Biophys. 167, 452–457 (1975).

    Article  PubMed  CAS  Google Scholar 

  • Parpinello G., Berardi E., Strabbioli R.: A regulatory mutant ofHansenula polymorpha exhibiting methanol utilization metabolism and peroxisome proliferation in glucose.J. Bacteriol. 180, 2958–2967 (1998).

    PubMed  CAS  Google Scholar 

  • Petit T., Gancedo C.: Molecular cloning and characterization of the geneHXK1 encoding the hexokinase fromYarrowia lipolytica.Yeast 15, 1573–1584 (1999).

    Article  PubMed  CAS  Google Scholar 

  • Prior C., Mamessier P., Fukuhara H., Chen X.J.: The hexokinase gene is required for transcriptional regulation of the glucose transporter geneRAG1 inKluyveromyces lactis.Mol. Cell. Biol. 13, 3882–3889 (1993).

    PubMed  CAS  Google Scholar 

  • Randez-Gil F., Herrero P., Sanz P., Prieto J.A., Moreno F.: Hexokinase PII has a double cytosolic-nuclear localization inSaccharomyces cerevisiae.FEBS Lett. 425, 475–478 (1998).

    Article  PubMed  CAS  Google Scholar 

  • Roggenkamp R.: Constitutive appearance of peroxisomes in a regulatory mutant of the methylotrophic yeastHansenula polymorpha.Mol. Gen. Genet. 213, 535–540 (1988).

    Article  PubMed  CAS  Google Scholar 

  • Roggenkamp R., Hansen H., Eckart M., Janowicz Z., Hollenberg C.P.: Transformation of the methylotrophic yeastHansenula polymorpha by autonomous replication and integration vectors.Mol. Gen. Genet. 202, 302–308 (1986).

    Article  CAS  Google Scholar 

  • Ronne H.: Glucose repression in fungi.Trends Genet. 11, 12–17 (1995).

    Article  PubMed  CAS  Google Scholar 

  • Rose M.: Molecular and biochemical characterization of the hexokinase from the starch-utilizing yeastSchwanniomyces occidentalis.Curr. Genet. 27, 330–338 (1995).

    Article  PubMed  CAS  Google Scholar 

  • Rose M., Albig W., Entian K.-D.: Glucose repression inSaccharomyces cerevisiae is directly associated with hexose phosphorylation by hexokinases PI and PII.Eur. J. Biochem. 199, 511–518 (1991).

    Article  PubMed  CAS  Google Scholar 

  • Ruijter G.J., Panneman H., van den Broeck H.C., Bennett J.M., Visser J.: Characterization of theAspergillus nidulans frA1 mutant: hexose phosphorylation and apparent lack of involvement of hexokinase in glucose repression.FEMS Microbiol. Lett. 139, 223–228 (1996).

    PubMed  CAS  Google Scholar 

  • Sakai Y., Sawai T., Tani Y.: Isolation and characterization of a catabolite repression-insensitive mutant of a methanol yeastCandida boidinii A5 producing alcohol oxidase in glucose-containing medium.Appl. Environ. Microbiol. 53, 1812–1818 (1987).

    PubMed  CAS  Google Scholar 

  • Sibirny A.A., Titorenko V.I., Gonchar M. V., Ubiyvovk V.M., Ksheminskaya G.P., Vitvitskaya O.P.: Genetic control of methanol utilization in yeasts.J. Basic Microbiol. 28, 293–319 (1988).

    Article  PubMed  CAS  Google Scholar 

  • Veale R.A., Giuseppin M.L.F., van Eijk H.M.J., Sudbery P.E., Verrips C.T.: Development of a strain ofHansenula polymorpha for the efficient expression of guar α-galactosidase.Yeast 8, 361–372 (1992).

    Article  PubMed  CAS  Google Scholar 

  • Walsh R.B., Clifton D., Horak J., Fraenkel D.G.:Saccharomyces cerevisiae null mutants in glucose phosphorylation: metabolism and invertase expression.Genetics 128, 521–527 (1991).

    PubMed  CAS  Google Scholar 

  • Wedlock D.N., Thornton R.J.: A hexokinase associated with catabolite repression inPachysolen tannophilus.J. Gen. Microbiol. 135, 2013–2018 (1989).

    CAS  Google Scholar 

  • Wedlock D.N., James A.P., Thornton R.J.: Glucose-negative mutants ofPachysolen tannophilus.J. Gen. Microbiol. 135, 2019–2026 (1989).

    CAS  Google Scholar 

  • Zimmermann F.K., Sheel I.: Mutants ofSaccharomyces cerevisiae resistant to catabolite repression.Mol. Gen. Genet. 154, 75–82 (1977).

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Genetics, Institute of Molecular and Cell Biology, University of Tartu, 51010, Tartu, Estonia

    T. Kramarenko, H. Karp, A. Järviste & T. Alamäe

Authors
  1. T. Kramarenko
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. H. Karp
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. A. Järviste
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. T. Alamäe
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to T. Alamäe.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kramarenko, T., Karp, H., Järviste, A. et al. Sugar repression in the methylotrophic yeastHansenula polymorpha studied by using hexokinase-negative, glucokinase-negative and double kinase-negative mutants. Folia Microbiol 45, 521–529 (2000). https://doi.org/10.1007/BF02818721

Download citation

  • Received:

  • Revised:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF02818721

Keywords

Search

Navigation