Malate transport in Schizosaccharomyces pombe

Osothsilp, C.; Subden, R. E.

doi:10.1128/jb.168.3.1439-1443.1986

Cited by 49 publications

(34 citation statements)

References 23 publications

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“…It has been suggested that the S. pombe L-malic acid transporter is a carboxylate/proton symport system, dependent on the plasma membrane proton gradient (22). The MAE1 gene, encoding the permease for L-malic acid and other organic acids, has been cloned (18).…”

Section: Discussionmentioning

confidence: 99%

“…The transport systems involved are inducible and subject to glucose repression. In contrast, it has been suggested that the uptake of L-malic acid is mediated by a protein and occurs in the presence of glucose in Zygosaccharomyces bailii (3) and in Schizosaccharomyces pombe (22). The heter-ologous expression of a gene encoding such an L-malic acid permease appeared to be a possible way of improving malate uptake in S. cerevisiae during alcoholic fermentation.…”

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confidence: 91%

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Characterization ofSchizosaccharomyces pombeMalate Permease by Expression inSaccharomyces cerevisiae

Camarasa

Bidard

Bony

et al. 2001

Appl Environ Microbiol

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In Saccharomyces cerevisiae, L-malic acid transport is not carrier mediated and is limited to slow, simple diffusion of the undissociated acid. Expression in S. cerevisiae of the MAE1 gene, encoding Schizosaccharomyces pombe malate permease, markedly increased L-malic acid uptake in this yeast. In this strain, at pH 3.5 (encountered in industrial processes), L-malic acid uptake involves Mae1p-mediated transport of the monoanionic form of the acid (apparent kinetic parameters: V max ‫؍‬ 8.7 nmol/mg/min; K m ‫؍‬ 1.6 mM) and some simple diffusion of the undissociated L-malic acid (K d ‫؍‬ 0.057 min ؊1 ). As total L-malic acid transport involved only low levels of diffusion, the Mae1p permease was further characterized in the recombinant strain. L-Malic acid transport was reversible and accumulative and depended on both the transmembrane gradient of the monoanionic acid form and the ⌬pH component of the proton motive force. Dicarboxylic acids with stearic occupation closely related to L-malic acid, such as maleic, oxaloacetic, malonic, succinic and fumaric acids, inhibited L-malic acid uptake, suggesting that these compounds use the same carrier. We found that increasing external pH directly inhibited malate uptake, resulting in a lower initial rate of uptake and a lower level of substrate accumulation. In S. pombe, proton movements, as shown by internal acidification, accompanied malate uptake, consistent with the proton/dicarboxylate mechanism previously proposed. Surprisingly, no proton fluxes were observed during Mae1p-mediated L-malic acid import in S. cerevisiae, and intracellular pH remained constant. This suggests that, in S. cerevisiae, either there is a proton counterflow or the Mae1p permease functions differently from a proton/dicarboxylate symport.Previous studies have provided evidence that L-malic acid is metabolized by Saccharomyces cerevisiae only in the presence of an assimilable carbon source (16,24,25). Exogenous L-malic acid (3 g/liter) is always consumed to a limited extent (10 to 20%), and the amount of degraded malate depends on the strain and culture conditions. This incomplete consumption of L-malic acid may be due to limited malate uptake and inefficiency of the enzyme systems involved in metabolism of the acid. Indeed, it has been reported that the transport of Lmalate is not carrier mediated in S. cerevisiae; the undissociated form of the acid slowly enters the cell by simple diffusion (28). During fermentation in the presence of malate, intracellular malate concentration in this yeast (close to 1 mM) is therefore lower than that in yeasts having a carrier protein for L-malate and able to metabolize L-malic acid completely (10 to 15 mM in Saccharomyces bailii) (20). The constitutive L-malic enzyme, thought to be responsible for the anaerobic metabolism of L-malate in S. cerevisiae, has a high K m for the substrate (50 mM) (16,17). Given the low levels of intracellular malate, the malic enzyme seems to function to a limited extent. Anaerobically grown S. cerevisiae cells also contain a ma...

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Section: Discussionmentioning

confidence: 99%

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confidence: 91%

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Characterization ofSchizosaccharomyces pombeMalate Permease by Expression inSaccharomyces cerevisiae

Camarasa

Bidard

Bony

et al. 2001

Appl Environ Microbiol

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“…Uptake of dicarboxylic acids in yeasts has been reported in several papers (4,9,11,12,26,28,32). It is believed that several yeast strains, such as Schizosaccharomyces pombe, Candida sphaerica, Candida utilis, and Hansenula anomala, can import dicarboxylic acids via a proton symport mechanism.…”

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pH-Dependent Uptake of Fumaric Acid in Saccharomyces cerevisiae under Anaerobic Conditions

Jamalzadeh

Verheijen

Heijnen

et al. 2012

Appl Environ Microbiol

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Microbial production of C 4 dicarboxylic acids from renewable resources has gained renewed interest. The yeast Saccharomyces cerevisiae is known as a robust microorganism and is able to grow at low pH, which makes it a suitable candidate for biological production of organic acids. However, a successful metabolic engineering approach for overproduction of organic acids requires an incorporation of a proper exporter to increase the productivity. Moreover, low-pH fermentations, which are desirable for facilitating the downstream processing, may cause back diffusion of the undissociated acid into the cells with simultaneous active export, thereby creating an ATP-dissipating futile cycle. In this work, we have studied the uptake of fumaric acid in S. cerevisiae in carbon-limited chemostat cultures under anaerobic conditions. The effect of the presence of fumaric acid at different pH values (3 to 5) has been investigated in order to obtain more knowledge about possible uptake mechanisms. The experimental results showed that at a cultivation pH of 5.0 and an external fumaric acid concentration of approximately 0.8 mmol · liter ؊1 , the fumaric acid uptake rate was unexpectedly high and could not be explained by diffusion of the undissociated form across the plasma membrane alone. This could indicate the presence of protein-mediated import. At decreasing pH levels, the fumaric acid uptake rate was found to increase asymptotically to a maximum level. Although this observation is in accordance with proteinmediated import, the presence of a metabolic bottleneck for fumaric acid conversion under anaerobic conditions could not be excluded.

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“…Cells of S. pombe are not able to utilize malate as the sole energy source or incorporate the malate into biomass (2) and therefore require glucose or other carbon sources for the energy-dependent transport and efficient degradation of malic acid (3). Three enzymes are involved in malate degradation in S. pombe, namely the malate transporter, malic enzyme, and malate dehydrogenase (4). The transporter, encoded by the mae1 gene (5), uses an H ϩ -symport system for the active transport of L-malate, and the NAD-dependent malic enzyme (EC 1.1.1.38) catalyzes the oxidative decarboxylation of L-malate to pyruvate and CO 2 .…”

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Transcriptional Regulation of the Schizosaccharomyces pombe Malic Enzyme Gene, mae2

Viljoen

Volschenk

Young

et al. 1999

Journal of Biological Chemistry

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The NAD-dependent malic enzyme from Schizosaccharomyces pombe catalyzes the oxidative decarboxylation of L-malate to pyruvate and CO 2 . Transcription of the S. pombe malic enzyme gene, mae2, was studied to elucidate the regulatory mechanisms involved in the expression of the gene. No evidence for substrate-induced expression of mae2 was observed in the presence of 0.2% L-malate. However, transcription of mae2 was induced when cells were grown in high concentrations of glucose or under anaerobic conditions. The increased levels of malic enzyme may provide additional pyruvate or assist in maintaining the redox potential under fermentative conditions. Deletion and mutation analyses of the 5-flanking region of the mae2 gene revealed the presence of three novel negative cis-acting elements, URS1, URS2, and URS3, that seem to function cooperatively to repress transcription of the mae2 gene. URS1 and URS2 are also present in the promoter region of the S. pombe malate transporter gene, suggesting co-regulation of their expression. Furthermore, two positive cisacting elements in the mae2 promoter, UAS1 and UAS2, show homology with the DNA recognition sites of the cAMP-dependent transcription factors ADR1, AP-2, and ATF (activating transcription factor)/CREB (cAMP response element binding).The fission yeast Schizosaccharomyces pombe efficiently degrades L-malate to CO 2 under aerobic conditions and to ethanol and CO 2 under anaerobic conditions (1). Cells of S. pombe are not able to utilize malate as the sole energy source or incorporate the malate into biomass (2) and therefore require glucose or other carbon sources for the energy-dependent transport and efficient degradation of malic acid (3). Three enzymes are involved in malate degradation in S. pombe, namely the malate transporter, malic enzyme, and malate dehydrogenase (4). The transporter, encoded by the mae1 gene (5), uses an H ϩ -symport system for the active transport of L-malate, and the NAD-dependent malic enzyme (EC 1.1.1.38) catalyzes the oxidative decarboxylation of L-malate to pyruvate and CO 2 . The mitochondrial malate dehydrogenase oxidizes L-malate to oxaloacetate in the tricarboxylic acid cycle and is responsible for 10% of the degradation of malate under aerobic conditions. Molecular analysis of the S. pombe malic mae2 gene showed a high degree of homology with malic enzymes from various organisms (6). Eight highly conserved regions were identified in malic enzymes, including the binding sites for L-malate and the dinucleotide co-factors NAD(P) ϩ (7,8). Although the secondary structure of malic enzymes is highly conserved, the coenzyme specificity (NAD ϩ or NADP ϩ ) and cellular localization (cytosolic or mitochondrial) are strongly linked to their regulation and metabolic function (9). Cytosolic NADP-dependent malic enzymes play an important role in lipid metabolism in higher eukaryotes (10), whereas NAD-dependent malic enzymes provide mitochondrial NADH for electron transport or cytosolic NADH for reductive power for other metabolic pathways (9)....

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Malate transport in Schizosaccharomyces pombe

Cited by 49 publications

References 23 publications

Characterization ofSchizosaccharomyces pombeMalate Permease by Expression inSaccharomyces cerevisiae

Characterization ofSchizosaccharomyces pombeMalate Permease by Expression inSaccharomyces cerevisiae

pH-Dependent Uptake of Fumaric Acid in Saccharomyces cerevisiae under Anaerobic Conditions

Transcriptional Regulation of the Schizosaccharomyces pombe Malic Enzyme Gene, mae2

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