1887

Abstract

This is a study of high-affinity glucose uptake in and the effect of disruption of a high-affinity monosaccharide-transporter gene, . The substrate saturation constant ( ) of a reference strain was about 15 μM in glucose-limited chemostat culture. Disruption of resulted in a two- to fivefold reduction in affinity for glucose and led to expression of a low-affinity glucose transport gene, , at high dilution rate. The effect of disruption was more subtle at low and intermediate dilution rates, pointing to some degree of functional redundancy in the high-affinity uptake system of . The disruptant and a reference strain were cultivated in glucose-limited chemostat cultures at low, intermediate and high dilution rate (=0.07 h, 0.14 h and 0.20 h). Mycelium harvested from steady-state cultures was subjected to glucose uptake assays, and analysed for expression of and two other transporter genes, and . The capacity for glucose uptake ( ) of both strains was significantly reduced at low dilution rate. The glucose uptake assays revealed complex uptake kinetics. This impeded accurate determination of maximum specific uptake rates ( ) and apparent affinity constants () at intermediate and high dilution rate. Two high-affinity glucose transporter genes, and , were expressed at all three dilution rates in chemostat cultures, in contrast to batch culture, where only was expressed. Expression patterns of the three transporter genes suggested differential regulation and functionality of their products.

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2007-06-01
2024-03-28
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References

  1. Agger T., Spohr A. B., Carlsen M., Nielsen J. 1998; Growth and product formation of Aspergillus oryzae during submerged cultivations: verification of a morphologically structured model using fluorescent probes. Biotechnol Bioeng 57:321–329 [CrossRef]
    [Google Scholar]
  2. Bainbridge B. W., Bull A. T., Pirt S. J., Rowley B. I., Trinci A. P. J. 1971; Biochemical and structural changes in non-growing, maintained and autolysing cultures of Aspergillus nidulans. Trans Br Mycol Soc 56:371–385 [CrossRef]
    [Google Scholar]
  3. Bergmeyer H. U., Bernt E., Schmidt F., Stork H. 1974; d-Glucose determination with hexokinase and glucose-6-phosphate dehydrogenase. In Methods of Enzymatic Analysis 3 pp 1196–1201 Edited by Bergmeyer H. U. New York: Academic Press;
    [Google Scholar]
  4. Brown C. E., Romano A. H. 1969; Evidence against necessary phosphorylation during hexose transport in Aspergillus nidulans. J Bacteriol 100:1198–1203
    [Google Scholar]
  5. Carlsen M., Nielsen J. B., Villadsen J. 1996; Growth and α -amylase production by Aspergillus oryzae during continuous cultivations. J Biotechnol 45:81–93 [CrossRef]
    [Google Scholar]
  6. Diderich J. A., Schepper M., Luttik A. H., Pronk J. T., Klaassen P., Boelens H. F. M. other authors van Hoek P., van Dijken J. P., de Mattos M. J. T. 1999; Glucose uptake kinetics and transcription of hxt genes in chemostat cultures of Saccharomyces cerevisiae. J Biol Chem 274:15350–15359 [CrossRef]
    [Google Scholar]
  7. du Preez J. C., de Kock S. H., Kilian S. G., Litthauer D. 2000; The relationship between transport kinetics and glucose uptake by Saccharomyces cerevisiae in aerobic chemostat cultures. Antonie Van Leeuwenhoek 77:379–388 [CrossRef]
    [Google Scholar]
  8. Fan J., Chaturvedi V., Shen S. 2002; Identification and phylogenetic analysis of a glucose transporter family from the human pathogenic yeast Candida albicans. J Mol Evol 55:336–346 [CrossRef]
    [Google Scholar]
  9. Fuhrmann G. F., Sander S., Potthast M., Völker B. 1989; Kinetic analysis and simulation of glucose transport in plasma membrane vesicles of glucose-repressed and derepressed Saccharomyces cerevisiae cells. Experientia 45:1018–1023 [CrossRef]
    [Google Scholar]
  10. Iversen J. J. L. 1981; A rapid sampling valve with minimal dead space for laboratory scale fermenters. Biotechnol Bioeng 23:437–440 [CrossRef]
    [Google Scholar]
  11. Larsen B., Rask Poulsen B., Eriksen N. T., Lønsmann Iversen J. J. 2004; Homogeneous batch cultures of Aspergillus oryzae by elimination of wall growth in the Variomixing bioreactor. Appl Microbiol Biotechnol 64:192–198 [CrossRef]
    [Google Scholar]
  12. MacCabe A. P., Ventura L., Miró P., Ramón D. 2003; Glucose uptake in germinating Aspergillus nidulans conidia: involvement of the creA and sorA genes. Microbiology 149:2129–2136 [CrossRef]
    [Google Scholar]
  13. Mackereth F. J. H. 1964; An improved galvanic cell for determination of oxygen concentrations in fluids. J Sci Instrum 41:38–41 [CrossRef]
    [Google Scholar]
  14. Mark C. G., Romano A. H. 1971; Properties of the hexose transport systems of Aspergillus nidulans. Biochim Biophys Acta 249:216–226 [CrossRef]
    [Google Scholar]
  15. Melchers W. J. G., Verweij P. E., Hoogkamp-Korstanje A. A., Meis J. F. G. M., van den Hurk P., van Belkum A., de Pauw B. E. 1994; General primer-mediated PCR for detection of Aspergillus species. J Clin Microbiol 32:1710–1717
    [Google Scholar]
  16. Mischak H., Kubicek C. P., Röhr M. 1984; Citrate inhibition of glucose uptake in Aspergillus niger. Biotechnol Lett 6:425–430 [CrossRef]
    [Google Scholar]
  17. Monod J. 1942 Recherches sur la Croissance des Cultures Bactériennes Paris: Hermann; (1958
    [Google Scholar]
  18. Monod J. 1950; La technique de culture continue: théorie et applications. Ann Inst Pasteur 79:390–410
    [Google Scholar]
  19. Moore D., Devadatham M. S. 1979; Sugar transport in Coprinus cinereus. Biochim Biophys Acta 550:515–526 [CrossRef]
    [Google Scholar]
  20. Pao S. S., Paulsen I. T., Saier M. H. 1998; Major facilitator superfamily. Microbiol Mol Biol Rev 62:1–34
    [Google Scholar]
  21. Peinado J. M., Cameira-Dos-Santos P. J., Loureiro-Días M. C. 1989; Regulation of glucose transport in Candida utilis. J Gen Microbiol 135:195–201
    [Google Scholar]
  22. Pirt S. J. 1975 Principles of Microbe and Cell Cultivation Oxford: Blackwell Scientific Publications;
    [Google Scholar]
  23. Pontecorvo G. 1953; The genetics of Aspergillus nidulans . In Advances in Genetics vol. 5 pp 141–238 Edited by Demerec M. New York: Academic Press;
    [Google Scholar]
  24. Postma E., Scheffers W. A., van Dijken J. P. 1988; Adaptation of the kinetics of glucose transport to environmental conditions in the yeast Candida utilis CBS 621: a continuous-culture study. J Gen Microbiol 134:1109–1116
    [Google Scholar]
  25. Postma E., Kuiper A., Tomasouw W. F., Scheffers A., van Dijken J. P. 1989a; Competition for glucose between the yeasts Saccharomyces cerevisiae and Candida utilis. Appl Environ Microbiol 55:3214–3220
    [Google Scholar]
  26. Postma E., Scheffers W. A., van Dijken J. P. 1989b; Kinetics of growth and glucose transport in glucose-limited chemostat cultures of Saccharomyces cerevisiae CBS 8066. Yeast 5:159–165 [CrossRef]
    [Google Scholar]
  27. Poulsen B. R., Iversen J. J. L. 1997; Mixing determinations in reactor vessels using linear buffers. Chem Eng Sci 52:979–984 [CrossRef]
    [Google Scholar]
  28. Rolland F., Winderickx J., Thevelein J. M. 2002; Glucose-sensing and -signalling mechanisms in yeast. FEMS Yeast Res 2:183–201 [CrossRef]
    [Google Scholar]
  29. Scarborough G. A. 1970a; Sugar transport in Neurospora crassa. J Biol Chem 245:1694–1698
    [Google Scholar]
  30. Scarborough G. A. 1970b; Sugar transport in Neurospora crassa . II. A second glucose transport system. J Biol Chem 245:3985–3987
    [Google Scholar]
  31. Torres N. V., Riol-Cimas J. M., Wolschek M., Kubicek C. P. 1996; Glucose transport by Aspergillus niger : the low-affinity carrier is only formed during growth on high glucose concentrations. Appl Microbiol Biotechnol 44:790–794
    [Google Scholar]
  32. vanKuyk P. A., Diderich J. A., MacCabe A. P., Hererro O., Ruijter G. J. G., Visser J. 2004; Aspergillus niger mstA encodes a high affinity sugar : H+ symporter which is regulated in response to extracellular pH. Biochem J 379:375–383 [CrossRef]
    [Google Scholar]
  33. Vishniac W., Santer M. 1957; The thiobacilli. Bacteriol Rev 21:195–213
    [Google Scholar]
  34. Walsh M. C., Smits H. P., Scholte M., van Dam K. 1994; Affinity of glucose transport in Saccharomyces cerevisiae is modulated during growth on glucose. J Bacteriol 176:953–958
    [Google Scholar]
  35. Wei H., Vienken K., Weber R., Bunting S., Requena N., Fischer R. 2004; A putative high affinity hexose transporter, hxtA , of Aspergillus nidulans is induced in vegetative hyphae upon starvation and in ascogenous hyphae during cleistothecium formation. Fungal Genet Biol 41:148–156 [CrossRef]
    [Google Scholar]
  36. Wieczorke R., Krampe S., Weierstall T., Freidel K., Hollenberg C. P., Boles E. 1999; Concurrent knock-out of at least 20 transporter genes is required to block uptake of hexoses in Saccharomyces cerevisiae. FEBS Lett 464:123–128 [CrossRef]
    [Google Scholar]
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