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Background Trehalose is a non-reducing disaccharide highly conserved throughout evolution. In yeasts, trehalose hydrolysis is confined to the enzyme trehalase, an α-glucosidase specific for trehalose as sole substrate. Two kinds of trehalase activity exist in yeasts: neutral and acid enzymes. Scope of the review This review makes a comparative survey of the main biochemical and genetic parameters, regulatory systems, tridimensional structure and catalytic mechanism of the two yeast trehalases. Major conclusions The yeast neutral and acid trehalases display sharp differences in biochemical features (optimum pH, Mr or amino acid sequence) physiological roles, subcellular location (cytosol vs vacuoles or cell wall) and regulatory control (phosphorylation vs catabolite repression). However, their identical specificity for trehalose is based on the presence of an (α/α)6 toroid folding structure in the active centre and a catalytic mechanism of anomeric inversion. General significance This review expands our knowledge of the homology, functional features and catalytic mechanisms of α-glucosidases in yeasts. It provides a further analysis of the correlation between structures and predicted biological roles of macromolecules.

A double homozygous atc1Δ/atc1Δ/ntc1Δ/ntc1Δ mutant (atc1Δ/ntc1Δ KO) was constructed in the pathogen opportunistic yeast Candida parapsilosis by disruption of the two chromosomal alleles coding for NTC1 gene (encoding a neutral trehalase) in a Cpatc1Δ/atc1Δ background (atc1Δ KO strain, deficient in acid trehalase). The Cpatc1Δ/ntc1Δ KO mutant failed to counteract the inability of Cpatc1Δ cells to metabolize exogenous trehalose and showed a similar growth pattern on several monosaccharides and disaccharides. However, upon prolonged incubation in either rich medium (YPD) or nutrient-starved medium the viability of Cpatc1Δ cells exhibited a sensitive phenotype, which was augmented by further CpNTC1/NTC1 disruption. Furthermore, Cpatc1Δ/ntc1Δ KO cells had difficulty in resuming active growth in fresh YPD. This homozygous mutant also lacked any in vitro measurable trehalase activity, whether acid or neutral, suggesting that a single gene codes for each enzyme. By contrast, in Cpatc1Δ/ntc1Δ KO strain the resistance to oxidative and heat stress displayed by atc1Δ mutant was suppressed. Cpatc1Δ/ntc1Δ KO cells showed a significant decrease in virulence as well as in the capacity to form biofilms. These results point to a major role for acid trehalase (Atc1p) in the pathobiology of C. parapsilosis, whereas the activity of neutral trehalase can only partially counteract Atc1p deficiency. They also support the use of ATC1 and NTC1 genes as interesting antifungal targets.

Agro-food industrial wastes represent a serious environmental problem in producing countries. In addition to the traditional irrigation with wastewaters, application of solids to the land, and the extraction of valuable compounds for the pharmaceutical, cosmetics and food industries, a microbiological approach is also available. In this review, the most promising microbiological processes for the valorization of wine and oil by-products are presented. Recently, there has been an upsurge in the exploitation of the waste materials generated by the wine and olive oil industries. Treatments by physical and chemical methods are generally expensive, and they do not often provide complete solutions. Microbiological processes for the treatment of olive and grape residues have seen worldwide applications, and they are considered to be environmentally friendly, reliable and, in most cases, cost effective. Production of added-value products such as edible mushrooms, biofuels, organic acids, polymers and enzymes appear to be the new frontier in by-products valorization. Our study has been focused in these two major food industries in the Mediterranean area, engaged in wine and olive oil production.

Malolactic fermentation (MLF) is a secondary bioconversion developed in some wines involving malic acid decarboxylation. The induction of MLF in wine by cultures of free and immobilized Oenococcus oeni cells was investigated. This work reports on the effect of surface charges in the immobilization material, a recently described fibrous sponge, as well as the pH and the composition of the media where cells are suspended. A chemical treatment provided positive charge to the sponges (DE or DEAE) and gave the highest cell loadings and subsequent resistance to removal. Preculture media to grow the malolactic bacteria before the immobilization procedure were also evaluated. We have established favorable conditions for growth (Medium of Preculture), suspension solution (Tartrate-Phosphate Buffer), suspension pH (3.5-5.5) and immobilization matrix (DE or DEAE cellulose sponge) to induce MLF in red wine. The use of a semi-continuous system permitted a high-efficiency malic acid conversion by 2 x 10(9) cfu sponge(-)(1) in at least four subsequent batch fermentations.

The effect of malolactic fermentation (MLF) on the volatile composition of red wines was studied by inoculation with selected lactic acid bacteria. Four wines were inoculated with different Oenococcus oeni (syn. Leuconostoc oenos) strains, the major malolactic species found in wines, and one was inoculated with a Lactobacillus sp. strain. A non inoculated wine was also analyzed to act as a control. Malolactic fermentation and evolution of non volatile compounds were followed by HPLC and after the depletion of the malic acid present in wine the volatile compounds were extracted and analyzed by gas chromatography with flame ionization and mass spectrometry. Wines which had undergone the MLF showed a significant increment in total higher alcohols, esters and acids that are important in the sensory properties and quality of wine.

The YWP1 gene encoding a specific mycelial cell wall protein of Yarrowia lipolytica has been cloned and expressed in Saccharomyces cerevisiae using different episomal plasmids. Because the plasmids pYAE35BB and pYAE35ES carrying the YWP1 gene (including the 5′ noncoding promoter sequences) failed to express it, the YWP1 gene was cloned under the control of GAL/CYC or ACT S. cerevisiae promoters. A main band with an apparent molecular mass of 70 kDa was detected by immunoblotting in the cell wall fraction of transformants. Ywp1 processing and incorporation to the cell wall were similar in both Y. lipolytica and S. cerevisiae but not in its final localization in the cell wall. In Y. lipolytica Ywp1 is covalently bound to the cell wall (it is released only by Zymolyase digestion), whereas in S. cerevisiae it was not (it was released by boiling SDS solutions). These results suggest that the sequences involved in recognition, anchoring of a protein to the cell wall, or the catalytic activities implicated are different, at least for Ywp1, in Y. lipolytica and S. cerevisiae. Another possibility is that the target for attachment of Ywp1 is missing or cryptic in the cell wall of S. cerevisiae.