ABSTRACT
Yeast selection for the wine industry in Spain started in 1950 for the understanding of the microbial ecology, and for the selection of optimal strains to improve the performance of alcoholic fermentation and the overall wine quality. This process has been strongly developed over the last 30 years, firstly on Saccharomyces cerevisiae, and, lately, with intense activity on non-Saccharomyces. Several thousand yeast strains have been isolated, identified and tested to select those with better performance and/or specific technological properties. The present review proposes a global survey of this massive ex-situ preservation of eukaryotic microorganisms, a reservoir of biotechnological solutions for the wine sector, overviewing relevant screenings that led to the selection of strains from 12 genera and 22 species of oenological significance. In the first part, the attention goes to the selection programmes related to relevant wine-producing areas (i.e. Douro, Extremadura, Galicia, La Mancha and Uclés, Ribera del Duero, Rioja, Sherry area, and Valencia). In the second part, the focus shifted on specific non-Saccharomyces genera/species selected from different Spanish and Portuguese regions, exploited to enhance particular attributes of the wines. A fil rouge of the dissertation is the design of tailored biotechnological solutions for wines typical of given geographic areas.
Subject(s)
Vitis , Wine , Saccharomyces cerevisiae , Wine/analysis , Portugal , Fermentation , BiotechnologyABSTRACT
Schizosaccharomyces was initially considered as a spoilage yeast because of the production of undesirable metabolites such as acetic acid, hydrogen sulfide, or acetaldehyde, but it currently seems to be of great value in enology.o ced Nevertheless, Schizosaccharomyces can reduce all of the malic acid in must, leading to malolactic fermentation. Malolactic fermentation is a highly complicated process in enology and leads to a higher concentration of biogenic amines, so the use of Schizosaccharomyces pombe can be an excellent tool for assuring wine safety. Schizosaccharomyces also has much more potential than only reducing the malic acid content, such as increasing the level of pyruvic acid and thus the vinylphenolic pyranoanthocyanin content. Until now, few commercial strains have been available and little research on the selection of appropriate yeast strains with such potential has been conducted. In this study, selected and wild Sc. pombe strains were used along with a Saccharomyces cerevisiae strain to ferment red grape must. The results showed significant differences in several parameters including non-volatile and volatile compounds, anthocyanins, biogenic amines and sensory parameters.
Subject(s)
Fermentation , Food Safety , Malates/metabolism , Saccharomyces cerevisiae/metabolism , Schizosaccharomyces/metabolism , Wine/microbiology , Acetaldehyde/metabolism , Acetic Acid/metabolism , Anthocyanins/metabolism , Biogenic Amines/metabolism , Pyruvic Acid/metabolism , Vitis/metabolism , Volatile Organic Compounds/metabolism , Wine/analysisABSTRACT
Plant K(+) transporters of the HAK family belong to four rather divergent phylogenetic clusters, although most of the transporters belong to clusters I or II. A simple phylogenetic analysis of fungal and plant HAK transporters suggests that an original HAK gene duplicated even before fungi and plants diverged, generating transporters that at present fulfill different functions in the plant. The HvHAK1 transporter belongs to cluster I and mediates high-affinity K(+) uptake in barley roots, but no function is known for the cluster II transporter, HvHAK2, which is not functional in yeast. The function of HvHAK2 was investigated by constructing HvHAK1-HAK2 chimeric transporters, which were not functional even when they included only short fragments of HvHAK2. Then, amino acids characteristic of cluster II in the N terminus and in the first transmembrane domain were introduced into HvHAK1. All of these changes increased the Rb(+) K(m), introducing minimal changes in the Na(+) K(m), which suggested that HvHAK2 is a low-affinity, Na(+)-sensitive K(+) transporter. Using a K(+)-defective Escherichia coli mutant, we functionally expressed HvHAK2 and found that the predicted characteristics were correct, as well as discovering that the bacterial expression of HvHAK2 is functional at pH 5.5 but not at 7.5. We discuss whether HvHAK2 may be a tonoplast transporter effective for vacuolar K(+) depletion in K(+) starved plants.
Subject(s)
Cation Transport Proteins/chemistry , Plant Proteins/metabolism , Amino Acid Sequence , Arabinose/pharmacology , Biological Transport , Dose-Response Relationship, Drug , Escherichia coli/genetics , Escherichia coli/metabolism , Hordeum/metabolism , Hydrogen-Ion Concentration , Kinetics , Molecular Sequence Data , Mutagenesis , Mutation , Plasmids/metabolism , Potassium/metabolism , Potassium Chloride/pharmacology , Protein Structure, Tertiary , Sequence Homology, Amino Acid , Time Factors , VacuolesABSTRACT
We have cloned the gene encoding the TRK transporter of the soil yeast Schwanniomyces occidentalis and obtained the HAK1 trk1 delta and the hak1 delta TRK1 mutant strains. Analyses of the transport capacities of these mutants have shown that (i) the HAK1 and the TRK1 potassium transporters are the only transporters operating at low and medium K+ concentrations (< 1 mM); (ii) the HAK1 transporter is functional at low pH but fails at high pH; and (iii) the TRK1 transporter functions at neutral and high pH and fails at low pH. At neutral pH, both transporters are functional, but HAK1 is not expressed, except at very low K+ concentrations (< 50 microM) where HAK1 is very effective. TRK1 is also involved in the control of the membrane potential.
Subject(s)
Carrier Proteins/metabolism , Cation Transport Proteins , Membrane Proteins/metabolism , Potassium/metabolism , Saccharomyces cerevisiae Proteins , Saccharomycetales/metabolism , Amino Acid Sequence , Carrier Proteins/genetics , Fungal Proteins/genetics , Fungal Proteins/metabolism , Ion Transport , Membrane Proteins/genetics , Molecular Sequence Data , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Saccharomycetales/geneticsABSTRACT
Two genes isolated from Schwanniomyces occidentalis, ENA1 and ENA2, encode P-type ATPases highly homologous to the Na-ATPases of Saccharomyces cerevisiae and complement the Na+ sensitivity of an S. cerevisiae mutant strain lacking its own Na-ATPases. The expression of both ENA1 and ENA2 was highly dependent on a high external pH, but whereas a high pH was sufficient for the expression of ENA2, the expression of ENA1 required a high pH and the presence of Na+. Disruption of ENA1 rendered the cells less tolerant to Na+ than the wild-type strain and decreased their capacity for Na+ extrusion. Disruption of ENA2 did not affect Na+ tolerance, but decreased both the growth at high pH and K+ efflux. We discuss these results and propose that fungal Na-ATPases should be considered alkali cation ATPases. By sequence comparison, we found that fungal Na-ATPases form a homogeneous group that can be distinguished from other cation-pumping P-type ATPases, except from the cta3 Ca-ATPase of Schizosaccharomyces pombe.
Subject(s)
Adenosine Triphosphatases/metabolism , Ascomycota/enzymology , Cation Transport Proteins , Fungal Proteins/metabolism , Potassium/metabolism , Saccharomyces cerevisiae Proteins , Sodium/metabolism , Adenosine Triphosphatases/genetics , Amino Acid Sequence , Ascomycota/genetics , Fungal Proteins/genetics , Hydrogen-Ion Concentration , Molecular Sequence Data , Saccharomyces cerevisiae , Sequence Alignment , Sodium-Potassium-Exchanging ATPaseABSTRACT
The yeast Schwanniomyces occidentalis has a high-affinity K+ uptake system with a high concentrative capacity, which is able to deplete the external K+ to < 0.03 microM. We have cloned the gene HAK1 of S.occidentalis which complements defective K+ uptake by trk1 and trk1 trk2 mutants of Saccharomyces cerevisiae. When HAK1 was expressed in a trk1 trk2 S.cerevisiae mutant, transport affinities for K+ and other alkali cations resembled those of S.occidentalis. The predicted amino acid sequence of the HAK1 protein shows significant homology with the hydrophobic region of the Kup transporter of Escherichia coli. In S.occidentalis HAK1 expresses in K(+)-limiting conditions. Our data indicate that in K(+)-starved cells the system encoded by HAK1 is the major K+ transporter of S.occidentalis.
Subject(s)
Ascomycota/metabolism , Escherichia coli/metabolism , Genes, Fungal , Potassium/pharmacokinetics , Amino Acid Sequence , Ascomycota/drug effects , Ascomycota/genetics , Gene Expression Regulation, Fungal/drug effects , Genes, Fungal/drug effects , Glucose/pharmacology , Ion Transport/drug effects , Kinetics , Molecular Sequence Data , Mutation , Neomycin/pharmacology , Rubidium/pharmacokineticsABSTRACT
Na+ efflux and Na+ tolerance depend on a putative P-type ATPase encoded by the gene ENA1(PMR2) in Saccharomyces cerevisiae and on a putative Na+/H+ antiporter encoded by the gene sod2 in Schizosaccharomyces pombe. This report shows that a sod2::ura4 mutant of S. pombe transformed with the ENA1 gene of S. cerevisiae expressed the ENA1 protein, and recovered Na+ efflux and Na+ tolerance. The efflux of Na+ in the wild strain of S. pombe was sensitive to the transmembrane Na+ and H+ gradients, whereas in the sod2::ura4 mutant transformed with ENA1 it was independent of these gradients. The data give further support to the notion that ENA1 and sod2 encode Na+ transporters and not regulators of the process of Na+ export; they show also the physiological consequences of exporting Na+ through an Na(+)-ATPase or an Na+/H+ antiporter.
Subject(s)
Adenosine Triphosphatases/genetics , Cation Transport Proteins , Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae/enzymology , Schizosaccharomyces/genetics , Biological Transport , Cloning, Molecular , Hydrogen-Ion Concentration , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/growth & development , Saccharomyces cerevisiae/metabolism , Schizosaccharomyces/growth & development , Schizosaccharomyces/metabolism , Sodium/metabolism , Sodium-Potassium-Exchanging ATPaseABSTRACT
The ENA2 gene encoding a P-type ATPase involved in Na+ and Li+ effluxes in Saccharomyces cerevisiae has been isolated. The putative protein encoded by ENA2 differs only in thirteen amino acids from the protein encoded by ENA1/PMR2. However, ENA2 has a very low level of expression and for this reason did not confer significant Li+ tolerance on a Li+ sensitive strain. ENA1 and ENA2 are the first two units of a tandem array of four highly homologous genes with probably homologous functions.