Current Research on Flavor Compounds in Fermented Food Products

Recent advancements in the field of food science have spurred a surge of research focused on unraveling the intricate world of flavor compounds in fermented food products [...

The Smell of Synthetic Biology: Engineering Strategies for Aroma Compound Production in Yeast

Yeast—especially Saccharomyces cerevisiae—have long been a preferred workhorse for the production of numerous recombinant proteins and other metabolites. S. cerevisiae is a noteworthy aroma compound producer and has also been exploited to produce foreign bioflavour compounds. In the past few years, important strides have been made in unlocking the key elements in the biochemical pathways involved in the production of many aroma compounds. The expression of these biochemical pathways in yeast often involves the manipulation of the host strain to direct the flux towards certain precursors needed for the production of the given aroma compound. This review highlights recent advances in the bioengineering of yeast—including S. cerevisiae—to produce aroma compounds and bioflavours. To capitalise on recent advances in synthetic yeast genomics, this review presents yeast as a significant producer of bioflavours in a fresh context and proposes new directions for combining engineering and biology principles to improve the yield of targeted aroma compounds

Overexpression of native PSE1 and SOD1 in Saccharomyces cerevisiae improved heterologous cellulase secretion

Engineering cellulolytic ability into the yeast Saccharomyces cerevisiae to create an organism for consolidated bioprocessing (CBP) will require the simultaneous production and secretion of a number of heterologous cellulases. In addition, the generally low secretion titers achieved by this yeast will have to be overcome. To this end two native S. cerevisiae genes, PSE1 and SOD1, were individually overexpressed by placing each gene under the transcriptional control of the constitutive PGK1 promoter. The effect of these genes on heterologous protein secretion of three cellulases – an exoglucanase encoded by cel6A of Neocallimastix patriciarum, a β-glucosidase encoded by cel3A of Saccharomycopsis fibuligera and an endoglucanase encoded by cel7B of Trichoderma reesei was investigated by integrating the PGK1P/T–PSE1 and PGK1P/T–SOD1 cassettes into S. cerevisiae strains producing the relevant cellulases. Transformants were obtained that showed significantly higher secreted protein yield, with a resulting heterologous protein activity that ranged between 10% and 373% higher compared to the parental strains when grown in complex media. When both PSE1 and SOD1 were overexpressed in the yeast that produced Cel3A, a dramatic 447% increase in β-glucosidase activity was observed. This study shows that cellulase secretion in S. cerevisiae could be greatly improved with strain engineering. However, it also demonstrated that such strain engineering may have very enzyme specific effects as the induction of Cel3A secretion was far greater than that of the other cellulases investigated. Identifying cellulases amenable to expression in S. cerevisiae and engineering strains to maximize heterologous protein secretion may be imperative to creating optimal strains for CBP and may have wider implications for heterologous protein secretion in S. cerevisiae in general.7 page(s

Binding of NADP + triggers an open-to-closed transition in a mycobacterial FabG β-ketoacyl-ACP reductase

International audienc

Effect of Isomixing on Grape Must Fermentations of ATF1–Overexpressing Wine Yeast Strains

Speeding up grape must fermentation would be of great economic benefit. We subjected Saccharomyces cerevisiae VIN13 and two recombinant VIN13-strains expressing ATF1 alleles under two different promoters (either PGK1 or HXT7) to four styles of grape must fermentations; we then assessed the effect of constantly stirring a must fermentation (isomixing). The four different fermentation setups were as follows: isomixed, closed in an ANKOM Rf Gas productions system; isomixed, open in a stirred tall tube cylinder; static, closed constituting a conventional fermentation in a wine bottle equipped with an airlock and static; and static, open in a tall tube cylinder (without stirring). We report on major fermentation parameters and the volatile aroma compositions generated in the finished wines. The primary fermentations of the strains subjected to constant stirring finished after 7 days, whereas the static fermentations reached dryness after 19 days. The wines derived from isomixed fermentations produced approximately 0.7% less ethanol compared to the unstirred fermentations. The speed that the isomixed fermentation took to reach completion may provide an alternative to static fermentations in the preparation of base wines for sparkling wine production. The observed increase of volatiles of isomixed fermentations merits further investigation

Development of Genetic Modification Tools for Hanseniasporauvarum

Apiculate yeasts belonging to the genus Hanseniaspora are commonly isolated from viticultural settings and often dominate the initial stages of grape must fermentations. Although considered spoilage yeasts, they are now increasingly becoming the focus of research, with several whole-genome sequencing studies published in recent years. However, tools for their molecular genetic manipulation are still lacking. Here, we report the development of a tool for the genetic modification of Hanseniaspora uvarum. This was employed for the disruption of the HuATF1 gene, which encodes a putative alcohol acetyltransferase involved in acetate ester formation. We generated a synthetic marker gene consisting of the HuTEF1 promoter controlling a hygromycin resistance open reading frame (ORF). This new marker gene was used in disruption cassettes containing long-flanking (1000 bp) homology regions to the target locus. By increasing the antibiotic concentration, transformants were obtained in which both alleles of the putative HuATF1 gene were deleted in a diploid H. uvarum strain. Phenotypic characterisation including fermentation in Müller-Thurgau must showed that the null mutant produced significantly less acetate ester, particularly ethyl acetate. This study marks the first steps in the development of gene modification tools and paves the road for functional gene analyses of this yeast

Characterization of a mycobacterial cellulase and its impact on biofilm- and drug-induced cellulose production

International audienceIt was recently shown that Mycobacterium tuberculosis produces cellulose which forms an integral part of its extracellular polymeric substances within a biofilm set-up. Using Mycobacterium smegmatis as a proxy model organism, we demonstrate that M. smegmatis biofilms treated with purified MSMEG\₆752 releases the main cellulose degradation-product (cellobiose), detected by using ionic chromatography, suggesting that MSMEG\₆752 encodes a cellulase. Its overexpression in M. smegmatis prevents spontaneous biofilm formation. Moreover, the method reported here allowed detecting cellobiose when M. smegmatis cultures were exposed to a subinhibitory dose of rifampicin. Overall, this study highlights the role of the MSMEG\₆752 in managing cellulose production induced during biofilm formation and antibiotic stress response

Aroma Profiles of Vitis vinifera L. cv. Gewürztraminer Must Fermented with Co-Cultures of Saccharomyces cerevisiae and Seven Hanseniaspora spp.

In this study, the aroma-production profiles of seven different Hanseniaspora strains, namely H. guilliermondii, H. meyeri, H. nectarophila, H. occidentalis, H. opuntiae, H. osmophila and H. uvarum were determined in a simultaneous co-inoculation with the wine yeast Saccharomyces cerevisiae Champagne Epernay Geisenheim (Uvaferm CEG). All co-inoculated fermentations with Hanseniaspora showed a dramatic increase in ethyl acetate levels except the two (H. occidentalis and H. osmophila) that belong to the so-called slow-evolving clade, which had no meaningful difference, compared to the S. cerevisiae control. Other striking observations were the almost complete depletion of lactic acid in mixed-culture fermentations with H. osmophila, the more than 3.7 mg/L production of isoamyl acetate with H. guilliermondii, the significantly lower levels of glycerol with H. occidentalis and the increase in certain terpenols, such as citronellol with H. opuntiae. This work allows for the direct comparison of wines made with different Hanseniapora spp. showcasing their oenological potential, including two (H. meyeri and H. nectarophila) previously unexplored in winemaking experiments

The Use of <i>Hanseniaspora occidentalis</i> in a Sequential Must Inoculation to Reduce the Malic Acid Content of Wine

In this study, the impact of the apiculate yeast Hanseniaspora occidentalis as a co-partner with Saccharomyces cerevisiae was investigated in a sequential-type mixed-culture fermentation of Muscaris grape must. As with other fermentation trials using Hanseniaspora strains, a significant increase in ethyl acetate was observed, but most intriguing was the almost complete abolition of malic acid (from 2.0 g/L to 0.1 g/L) in the wine. Compared to the pure S. cerevisiae inoculum, there was also a marked increase in the concentrations of the other acetate esters. Modulation of some of the varietal elements, such as rose oxide, was also observed. This work shows the promising use of H. occidentalis in a mixed-culture must fermentation, especially in the acid modulation of fruit juice matrices