Tags

Type your tag names separated by a space and hit enter

Physiological characterisation of a pyruvate-carboxylase-negative Saccharomyces cerevisiae mutant in batch and chemostat cultures.
Antonie Van Leeuwenhoek. 1998 Nov; 74(4):253-63.AV

Abstract

A prototrophic pyruvate-carboxylase-negative (Pyc-) mutant was constructed by deleting the PYC1 and PYC2 genes in a CEN.PK strain of Saccharomyces cerevisiae. Its maximum specific growth rate on ethanol was identical to that of the isogenic wild type but it was unable to grow in batch cultures in glucose-ammonia media. Consistent with earlier reports, growth on glucose could be restored by supplying aspartate as a sole nitrogen source. Ethanol could not replace aspartate as a source of oxaloacetate in batch cultures. To investigate whether alleviation of glucose repression allowed expression of alternative pathways for oxaloacetate synthesis, the Pyc- strain and an isogenic wild-type strain were grown in aerobic carbon-limited chemostat cultures at a dilution rate of 0.10 h-1 on mixtures of glucose and ethanol. In such mixed-substrate chemostat cultures of the Pyc- strain, steady-state growth could only be obtained when ethanol contributed 30% or more of the substrate carbon in the feed. Attempts to further decrease the ethanol content of the feed invariably resulted in washout. In Pyc- as well as in wild-type cultures, levels of isocitrate lyase, malate synthase and phospho-enol-pyruvate carboxykinase in cell extracts decreased with a decreasing ethanol content in the feed. Nevertheless, at the lowest ethanol fraction that supported growth of the Pyc- mutant, activities of the glyoxylate cycle enzymes in cell extracts were still sufficient to meet the requirement for C4-compounds in biomass synthesis. This suggests that factors other than glucose repression of alternative routes for oxaloacetate synthesis prevent growth of Pyc- mutants on glucose.

Authors+Show Affiliations

de Jong-Gubbels P
Kluyver Institute of Biotechnology, Delft University of Technology, The Netherlands.
Bauer J
No affiliation info available
Niederberger P
No affiliation info available
Stückrath I
No affiliation info available
Kötter P
No affiliation info available
van Dijken JP
No affiliation info available
Pronk JT
No affiliation info available

MeSH

AmmoniaAspartic AcidCulture MediaEthanolGene Expression Regulation, FungalGlucoseGlyoxylatesMutationPhenotypePyruvate CarboxylaseSaccharomyces cerevisiae

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

10081585

Citation

de Jong-Gubbels, P, et al. "Physiological Characterisation of a Pyruvate-carboxylase-negative Saccharomyces Cerevisiae Mutant in Batch and Chemostat Cultures." Antonie Van Leeuwenhoek, vol. 74, no. 4, 1998, pp. 253-63.
de Jong-Gubbels P, Bauer J, Niederberger P, et al. Physiological characterisation of a pyruvate-carboxylase-negative Saccharomyces cerevisiae mutant in batch and chemostat cultures. Antonie Van Leeuwenhoek. 1998;74(4):253-63.
de Jong-Gubbels, P., Bauer, J., Niederberger, P., Stückrath, I., Kötter, P., van Dijken, J. P., & Pronk, J. T. (1998). Physiological characterisation of a pyruvate-carboxylase-negative Saccharomyces cerevisiae mutant in batch and chemostat cultures. Antonie Van Leeuwenhoek, 74(4), 253-63.
de Jong-Gubbels P, et al. Physiological Characterisation of a Pyruvate-carboxylase-negative Saccharomyces Cerevisiae Mutant in Batch and Chemostat Cultures. Antonie Van Leeuwenhoek. 1998;74(4):253-63. PubMed PMID: 10081585.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Physiological characterisation of a pyruvate-carboxylase-negative Saccharomyces cerevisiae mutant in batch and chemostat cultures. AU - de Jong-Gubbels,P, AU - Bauer,J, AU - Niederberger,P, AU - Stückrath,I, AU - Kötter,P, AU - van Dijken,J P, AU - Pronk,J T, PY - 1999/3/19/pubmed PY - 1999/3/19/medline PY - 1999/3/19/entrez SP - 253 EP - 63 JF - Antonie van Leeuwenhoek JO - Antonie Van Leeuwenhoek VL - 74 IS - 4 N2 - A prototrophic pyruvate-carboxylase-negative (Pyc-) mutant was constructed by deleting the PYC1 and PYC2 genes in a CEN.PK strain of Saccharomyces cerevisiae. Its maximum specific growth rate on ethanol was identical to that of the isogenic wild type but it was unable to grow in batch cultures in glucose-ammonia media. Consistent with earlier reports, growth on glucose could be restored by supplying aspartate as a sole nitrogen source. Ethanol could not replace aspartate as a source of oxaloacetate in batch cultures. To investigate whether alleviation of glucose repression allowed expression of alternative pathways for oxaloacetate synthesis, the Pyc- strain and an isogenic wild-type strain were grown in aerobic carbon-limited chemostat cultures at a dilution rate of 0.10 h-1 on mixtures of glucose and ethanol. In such mixed-substrate chemostat cultures of the Pyc- strain, steady-state growth could only be obtained when ethanol contributed 30% or more of the substrate carbon in the feed. Attempts to further decrease the ethanol content of the feed invariably resulted in washout. In Pyc- as well as in wild-type cultures, levels of isocitrate lyase, malate synthase and phospho-enol-pyruvate carboxykinase in cell extracts decreased with a decreasing ethanol content in the feed. Nevertheless, at the lowest ethanol fraction that supported growth of the Pyc- mutant, activities of the glyoxylate cycle enzymes in cell extracts were still sufficient to meet the requirement for C4-compounds in biomass synthesis. This suggests that factors other than glucose repression of alternative routes for oxaloacetate synthesis prevent growth of Pyc- mutants on glucose. SN - 0003-6072 UR - https://www.unboundmedicine.com/medline/citation/10081585/Physiological_characterisation_of_a_pyruvate_carboxylase_negative_Saccharomyces_cerevisiae_mutant_in_batch_and_chemostat_cultures_ DB - PRIME DP - Unbound Medicine ER -