Strains Of Yeast Saccharomyces Cerevisiae Research Articles

Studies on Toxicity Effect of Solvents on Growth of Saccharomyces cerevisiae (NCIM 3186)

The present study was conducted to determine the tolerance level of the yeast strain Saccharomyces cerevisiae (NCIM 3186) towards a few selected organic solvents, viz. ethyl oleate, ethyl benzene, ethyl caprylate and decanol (decyl alcohol). The objective of the investigation was to search for a suitable solvent with low toxicity level on yeast cells yet having high separation ability so that it can be used for simultaneous fermentation and extraction of ethanol in a future study. The result shows that ethyl oleate can be regarded as the best suited solvent in comparison to other solvents studied in the present investigation. The order of preference of a solvent which may be used for extractive fermentation is: ethyl oleate > ethyl benzene > ethyl caprylate > decanol (decyl alcohol).

Relationship of trehalose accumulation with ethanol fermentation in industrial Saccharomyces cerevisiae yeast strains

The protective effect and the mechanisms of trehalose accumulation in industrial Saccharomyces cerevisiae strains were investigated during ethanol fermentation. The engineered strains with more intercellular trehalose achieved significantly higher fermentation rates and ethanol yields than their wild strain ZS during very high gravity (VHG) fermentation, while their performances were not different during regular fermentation. The VHG fermentation performances of these strains were consistent with their growth capacity under osmotic stress and ethanol stress, the key stress factors during VHG fermentation. These results suggest that trehalose accumulation is more important for VHG fermentation of industrial yeast strains than regular one. The differences in membrane integrity and antioxidative capacity of these strains indicated the possible mechanisms of trehalose as a protectant under VHG condition. Therefore, trehalose metabolic engineering may be a useful strategy for improving the VHG fermentation performance of industrial yeast strains.

Alcoholic Fermentation with Flocculant Saccharomyces cerevisiae in Fed-Batch Process

Studies have been conducted on selecting yeast strains for use in fermentation for ethanol production to improve the performance of industrial plants and decrease production costs. In this paper, we study alcoholic fermentation in a fed-batch process using a Saccharomyces cerevisiae yeast strain with flocculant characteristics. Central composite design (CCD) was used to determine the optimal combination of the variables involved, with the sucrose concentration of 170 g/L, a cellular concentration in the inoculum of 40% (v/v), and a filling time of 6 h, which resulted in a 92.20% yield relative to the theoretical maximum yield, a productivity of 6.01 g/L h and a residual sucrose concentration of 44.33 g/L. With some changes in the process such as recirculation of medium during the fermentation process and increase in cellular concentration in the inoculum after use of the CCD was possible to reduce the residual sucrose concentration to 2.8 g/L in 9 h of fermentation and increase yield and productivity for 92.75% and 9.26 g/L h, respectively. A model was developed to describe the inhibition of alcoholic fermentation kinetics by the substrate and the product. The maximum specific growth rate was 0.103 h(-1), with K(I) and K(s) values of 109.86 and 30.24 g/L, respectively. The experimental results from the fed-batch reactor show a good fit with the proposed model, resulting in a maximum growth rate of 0.080 h(-1).

Abstract A151: Identification of antibody fragments specific for high-grade prostatic intraepithelial neoplasia cells via SELEX.

Abstract The purpose of this study is to develop a minimally invasive molecular tool for the early detection and specific treatment of prostate cancer. A human non-immune single-chain Fragment variable (scFv) library with 10∧9 diversity has been displayed by a yeast host surface with the ability to bind to whole cell targets. This protein library was cloned into plasmids and transformed into the yeast strain Saccharomyces cerevisiae. Surface expression of the scFv library was achieved by expressing the scFv under control of a galactose promoter preceding incubation with the target cell line. A molecular recognition element (MRE) specific for HGPIN cells will be selected for through the Systematic Evolution of Ligands by Exponential Enrichment (SELEX). From an initial scFv library with 10∧9 diversity, seven rounds of selection will be completed in order to enrich the antibody fragment library that binds only to the prostate cancer cell line HGPIN and not those that bind to normal and cancerous cells of the prostate epithelium. This involves screening a library of random molecules for their ability to bind to a target; non-binding molecules are removed and binding molecules are amplified. Amplified molecules are subjected to negative targets, and molecules that don't bind to these are amplified, thus completing a round of selection. Stringency is increased through each round of selection by shortening the yeast + prostate cell incubation period and decreasing the number of target cells used in incubation. After multiple rounds, one or a few strong binding, highly specific molecules are selected for. Diversity of the library is decreased with each round of selection. This is followed by amplification of the remaining yeast. Sequencing is performed throughout the selection process in order to monitor both the decrease in diversity and the emergence of the most selective sequences. The obtained MRE will be useful for specific diagnosis in the pre-malignant stages of prostate cancer. It may also be conjugated to drugs for targeted therapeutic applications. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A151. Citation Format: Cyrus J. Hajiran, Letha J. Sooter. Identification of antibody fragments specific for high-grade prostatic intraepithelial neoplasia cells via SELEX. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A151.

A Member of the Heavy Metal P-Type ATPase OsHMA5 Is Involved in Xylem Loading of Copper in Rice

Heavy metal-transporting P-type ATPase (HMA) has been implicated in the transport of heavy metals in plants. Here, we report the function and role of an uncharacterized member of HMA, OsHMA5 in rice (Oryza sativa). Knockout of OsHMA5 resulted in a decreased copper (Cu) concentration in the shoots but an increased Cu concentration in the roots at the vegetative stage. At the reproductive stage, the concentration of Cu in the brown rice was significantly lower in the mutants than in the wild-type rice; however, there was no difference in the concentrations of iron, manganese, and zinc between two independent mutants and the wild type. The Cu concentration of xylem sap was lower in the mutants than in the wild-type rice. OsHMA5 was mainly expressed in the roots at the vegetative stage but also in nodes, peduncle, rachis, and husk at the reproductive stage. The expression was up-regulated by excess Cu but not by the deficiency of Cu and other metals, including zinc, iron, and manganese, at the vegetative stage. Analysis of the transgenic rice carrying the OsHMA5 promoter fused with green fluorescent protein revealed that it was localized at the root pericycle cells and xylem region of diffuse vascular bundles in node I, vascular tissues of peduncle, rachis, and husk. Furthermore, immunostaining with an antibody against OsHMA5 revealed that it was localized to the plasma membrane. Expression of OsHMA5 in a Cu transport-defective mutant yeast (Saccharomyces cerevisiae) strain restored the growth. Taken together, OsHMA5 is involved in loading Cu to the xylem of the roots and other organs.

SILVER NANOPARTICLE IMPREGNATED BIO-BASED ACTIVATED CARBON WITH ENHANCED ANTIMICROBIAL ACTIVITY

The present study involves the production of silver nanoparticles using a novel yeast strain Saccharomyces cerevisiae BU-MBT CY-1 isolated from coconut cell sap. The biological reduction of silver nitrate by the isolate was deducted at various time intervals. The yeast cells after biological silver reduction were harvested and subjected to carbonization at 400°C for 1 h and its properties were analyzed using Fourier transform infra-red spectroscopy, X-ray diffraction, scanning electron microscope attached with energy dispersive spectroscopy and transmission electron microscopy. The average size of the silver nanoparticles present on the surface of the carbonized silver containing yeast cells (CSY) was 19 ± 9 nm. The carbonized control yeast cells (CCY) did not contain any particles on its surface. The carbonized silver nanoparticles containing yeast cells (CSY) were made into bioactive emulsion and tested for its efficacy against various pathogenic Gram positive and Gram negative bacteria. The antimicrobial activity studies indicated that CSY bioactive nanoemulsion was effective against Gram negative organisms than Gram positive organism.

Optimization of amorphadiene production in engineered yeast by response surface methodology.

Isoprenoids are among the most diverse bioactive compounds synthesized by biological systems. The superiority of these compounds has expanded their utility from pharmaceutical to fragrances, including biofuel industries. In the present study, an engineered yeast strain Saccharomyces cerevisiae (YCF-AD1) was optimized for production of Amorpha-4, 11-diene, a precursor of anti-malarial drug using response surface methodology. The effect of four critical parameters such as KH2PO4, methionine, pH and temperature were evaluated both qualitatively and quantitatively and further optimized for enhanced amorphadiene production by using a central composite design and model validation. The “goodness of fit” of the regression equation and model fit (R2) of 0.9896 demonstrate this study to be an effective model. Further, this model will be used to validate theoretically and experimentally at the higher level of amorphadiene production with the combination of the optimized values of KH2PO4 (4.0), methionine (1.49), pH (5.4) and temperature (33 °C).

A co-beneficial system using aquatic plants: bioethanol production from free-floating aquatic plants used for water purification

A co-beneficial system using constructed wetlands (CWs) planted with aquatic plants is proposed for bioethanol production and nutrient removal from wastewater. The potential for bioethanol production from aquatic plant biomass was experimentally evaluated. Water hyacinth and water lettuce were selected because of their high growth rates and easy harvestability attributable to their free-floating vegetation form. The alkaline/oxidative pretreatment was selected for improving enzymatic hydrolysis of the aquatic plants. Ethanol was produced with yields of 0.14-0.17 g-ethanol/ g-biomass in a simultaneous saccharification and fermentation mode using a recombinant Escherichia coli strain or a typical yeast strain Saccharomyces cerevisiae. Subsequently, the combined benefits of the CWs planted with the aquatic plants for bioethanol production and nutrient removal were theoretically estimated. For treating domestic wastewater at 1,100 m(3)/d, it was inferred that the anoxic-oxic activated sludge process consumes energy at 3,200 MJ/d, whereas the conventional activated sludge process followed by the CW consumes only 1,800 MJ/d with ethanol production at 115 MJ/d.

Xylosylation as an effective means for reducing yeast growth inhibition by 2‐phenylethanol

Microbial bioproduction processes of 2-phenylethanol, an important rose-like flavor and fragrance compound that occurs naturally in the essential oils of many flowers and plants, are hindered by the growth inhibition it exerts towards the producing microorganism, mainly yeast. We show here for the first time that glycosylation of 2-phenylethanol with xylose increased the inhibitory concentration inducing 50% decrease of the yeast Saccharomyces cerevisiae strain BY4741 growth rate (IC50 ) from 14 mM (1.71 g/L) 2-phenylethanol (2PE) to 100 mM (25.5 g/L) 2-phenylethyl β-D-xylopyranoside (X-2PE). More interestingly, the IC10 was only 3 mM (0.37 g/L) for 2PE and 86 mM (21.9 g/L) for X-2PE. Xylosylation of 2-phenylethanol can offer therefore an effective means for reducing yeast growth inhibition in microbial bioproduction processes of this important flavor and fragrance compound.

Dysfunctional mitochondria modulate cAMP-PKA signaling and filamentous and invasive growth of Saccharomyces cerevisiae.

Mitochondrial metabolism is targeted by conserved signaling pathways that mediate external information to the cell. However, less is known about whether mitochondrial dysfunction interferes with signaling and thereby modulates the cellular response to environmental changes. In this study, we analyzed defective filamentous and invasive growth of the yeast Saccharomyces cerevisiae strains that have a dysfunctional mitochondrial genome (rho mutants). We found that the morphogenetic defect of rho mutants was caused by specific downregulation of FLO11, the adhesin essential for invasive and filamentous growth, and did not result from general metabolic changes brought about by interorganellar retrograde signaling. Transcription of FLO11 is known to be regulated by several signaling pathways, including the filamentous-growth-specific MAPK and cAMP-activated protein kinase A (cAMP-PKA) pathways. Our analysis showed that the filamentous-growth-specific MAPK pathway retained functionality in respiratory-deficient yeast cells. In contrast, the cAMP-PKA pathway was downregulated, explaining also various phenotypic traits observed in rho mutants. Thus, our results indicate that dysfunctional mitochondria modulate the output of the conserved cAMP-PKA signaling pathway.

Performance evaluation of Pichia kluyveri, Kluyveromyces marxianus and Saccharomyces cerevisiae in industrial tequila fermentation

Traditionally, industrial tequila production has used spontaneous fermentation or Saccharomyces cerevisiae yeast strains. Despite the potential of non-Saccharomyces strains for alcoholic fermentation, few studies have been performed at industrial level with these yeasts. Therefore, in this work, Agave tequilana juice was fermented at an industrial level using two non-Saccharomyces yeasts (Pichia kluyveri and Kluyveromyces marxianus) with fermentation efficiency higher than 85 %. Pichia kluyveri (GRO3) was more efficient for alcohol and ethyl lactate production than S. cerevisiae (AR5), while Kluyveromyces marxianus (GRO6) produced more isobutanol and ethyl-acetate than S. cerevisiae (AR5). The level of volatile compounds at the end of fermentation was compared with the tequila standard regulation. All volatile compounds were within the allowed range except for methanol, which was higher for S. cerevisiae (AR5) and K. marxianus (GRO6). The variations in methanol may have been caused by the Agave tequilana used for the tests, since this compound is not synthesized by these yeasts.

The Kinetics of Alcoholic Fermentation by Two Yeast Strains in High Sugar Concentration Media

Over the last two decades, most of Italian vines have produced grapes with higher sugar to total acid ratios, greater concentrations of phenols and aromatic compounds and greater potential wine quality. As a consequence, the musts obtained by these grapes are more difficult to process because of the risk of slowing or stuck of fermentation. With the aim of describing the time evolution of the sugars bioconversion during alcoholic fermentation, the kinetics of the D-glucose and D-fructose degradations, promoted by two yeast strains (Saccharomyces cerevisiae (strain C) e Saccharomyces bayanus (strain B)), was investigated using synthetic media, added or not with ethanol. The concentrations of both the substrates and the products of the sugars conversions, as well as the number of viable cells of yeasts, were determined as a function of the alcoholic fermentation time and the related kinetics constants determined. If the reaction medium contained high concentrations of both glucose and fructose, the strains showed significant different fermentatory ability. In these conditions a stuck of fermentation occurred and the remaining sugar was only fructose (strain C) or prevailing fructose (strain B). If the reaction medium contained only glucose as substrate, the strain C seemed more efficient while the kinetics behavior changed completely in presence of only fructose. On the basis of the information collected using this kinetic approach, it would be possible to develop technical data sheets, specific for each yeast strain, useful to choose the optimal microbial strain as a function of the different operative conditions. Moreover the kinetic constant of hexose conversion could be adopted as bio-markers in selection and breeding of wine yeast strains having a lower tendency for sluggish fructose fermentation.

Combination ofERG9Repression and Enzyme Fusion Technology for Improved Production of Amorphadiene inSaccharomyces cerevisiae

The yeast strain (Saccharomyces cerevisiae) MTCC 3157 was selected for combinatorial biosynthesis of plant sesquiterpene amorpha-4,11-diene. Our main objective was to overproduce amorpha 4-11-diene, which is a key precursor molecule of artemisinin (antimalarial drug) produced naturally in plant Artemisia annua through mevalonate pathway. Farnesyl diphosphate (FPP) is a common intermediate metabolite of a variety of compounds in the mevalonate pathway of yeast and leads to the production of ergosterols, dolichol and ubiquinone, and so forth. In our studies, FPP converted to amorphadiene (AD) by expressing heterologous amorphadiene synthase (ADS) in yeast. First, ERG9 (squalane synthase) promoter of yeast was replaced with repressible methionine (MET3) promoter by using bipartite gene fusion method. Further to overcome the loss of the intermediate FPP through competitive pathways in yeast, fusion protein technology was adopted and farnesyldiphosphate synthase (FPPS) of yeast has been coupled with amorphadiene synthase (ADS) of plant origin (Artemisia annua L.) where amorphadiene production was improved by 2-fold (11.2 mg/L) and 4-fold (25.02 mg/L) in yeast strains YCF-002 and YCF-005 compared with control strain YCF-AD (5.5 mg/L), respectively.

The impact of different ale brewer’s yeast strains on the proteome of immature beer

BackgroundIt is well known that brewer’s yeast affects the taste and aroma of beer. However, the influence of brewer’s yeast on the protein composition of beer is currently unknown. In this study, changes of the proteome of immature beer, i.e. beer that has not been matured after fermentation, by ale brewer’s yeast strains with different abilities to degrade fermentable sugars were investigated.ResultsBeers were fermented from standard hopped wort (13° Plato) using two ale brewer’s yeast (Saccharomyces cerevisiae) strains with different attenuation degrees. Both immature beers had the same alcohol and protein concentrations. Immature beer and unfermented wort proteins were analysed by 2-DE and compared in order to determine protein changes arising from fermentation. Distinct protein spots in the beer and wort proteomes were identified using Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and MS/MS and revealed common beer proteins, such as lipid transfer proteins (LTP1 and LTP2), protein Z and amylase-protease inhibitors. During fermentation, two protein spots, corresponding to LTP2, disappeared, while three protein spots were exclusively found in beer. These three proteins, all derived from yeast, were identified as cell wall associated proteins, that is Exg1 (an exo-β-1,3-glucanase), Bgl2 (an endo-β-1,2-glucanase), and Uth1 (a cell wall biogenesis protein).ConclusionYeast strain dependent changes in the immature beer proteome were identified, i.e. Bgl2 was present in beer brewed with KVL011, while lacking in WLP001 beer.

39. Freezing effects evaluated on complex biological systems by DSC: Baker’s yeast and microalgal cells

When biomass is to be used in the food, feed, or pharmaceutical industry, it should be preserved and stored in order to permit transport from production to consumer. The most frequent preservation methods are freezing and drying. Differential scanning calorimetry (DSC) can be considered a powerful technique for studying thermal transitions of biological molecules related to their functionality and with biomass viability. Also, DSC has been shown to provide a convenient way to determine the chemical stability of some biomass components, such as lipids. The objective of this work was to analyze the feasibility to employ DSC as a tool to evaluate the cryo-resistance of baker’s industrial yeast ( Saccharomyces cerevisiae ) strain towards freezing and thawing and the long term conservation of intracellular lipids in Aurantiochytrium limacinum SR21 frozen microalgal cells. Yeast in stationary growth phase were exposed to two cycles of freezing (20 h and 4 h) at −20 °C after being exposed to various physical and chemical mild stresses. DSC runs were performed at 10 °C/min from 20 °C to 100 °C. Media with different C:N balances (55:1 and 6:1) were used to obtain microalgae with different PUFAs and carotenoid contents. Cells were frozen for 1, 8.5 and 30 months at −20 °C. The changes in the saturated/poly-unsaturated fatty acids (SFA/PUFAs) ratio in microalgal cells were detected by DSC under oxidative conditions (at 5 °C/min) through shifts in the onset oxidation temperature (OOT) values. Thermograms of the yeast’s systems showed two peaks that were attributed to low and high protein onset denaturation temperatures: 47–58 °C and 67–77 °C, respectively. Some pre-stress treatments (temperature, ethanol and NaCl) increased both enthalpy values and survival after freezing and thawing. DHA (C22:6, n − 3) and palmitic acid were the major PUFA and SFA, respectively produced by the SR21 cells. Microalgal growth carried out in media with 55:1 C:N ratio produced cells with higher levels of DHA and of carotenoids, when compared with those grown in media with a 6:1 C:N ratio. OOT of cells and of lipidic extracts were influenced by cellular carotenoids. Fatty acid stability in frozen microalgae could be evaluated by analyzing the shift of OOT values: the OOT values were lower for the frozen cells compared to the fresh ones and decreased as a function of frozen storage time. This behaviour could be associated with DHA degradation, in addition to carotenoid content lost. DSC onset denaturation temperatures clearly correlated with protein stability and the decrease in the enthalpy values obtained from the thermograms can provide valuable information about the corresponding effect on yeast viability. Besides, lipid stability in frozen media could be evaluated in whole microalgae cells as well as in their lipidic extracts analyzing the shift of OOT values. DSC has a high potential, as a non-chemical method, to determine stability parameters in complex biological systems and could be employed as an efficient, reliable and fast method to study proteins and fatty acids stability in whole cells and in their extracts. This work was supported by CONICET (PIP 100846, 100468), UBA (2002100100397 and I038) and ANPYCT (PICT 0928). Conflict of interest: None declared.

Dynamic study of yeast species and Saccharomyces cerevisiae strains during the spontaneous fermentations of Muscat blanc in Jingyang, China

The evolution of yeast species and Saccharomyces cerevisiae genotypes during spontaneous fermentations of Muscat blanc planted in 1957 in Jingyang region of China was followed in this study. Using a combination of colony morphology on Wallerstein Nutrient (WLN) medium, sequence analysis of the 26S rDNA D1/D2 domain and 5.8S-ITS-RFLP analysis, a total of 686 isolates were identified at the species level. The six species identified were S. cerevisiae, Hanseniaspora uvarum, Hanseniaspora opuntiae, Issatchenkia terricola, Pichia kudriavzevii (Issatchenkia orientalis) and Trichosporon coremiiforme. This is the first report of T. coremiiforme as an inhabitant of grape must. Three new colony morphologies on WLN medium and one new 5.8S-ITS-RFLP profile are described. Species of non-Saccharomyces, predominantly H. opuntiae, were found in early stages of fermentation. Subsequently, S. cerevisiae prevailed followed by large numbers of P. kudriavzevii that dominated at the end of fermentations. Six native genotypes of S. cerevisiae were determined by interdelta sequence analysis. Genotypes III and IV were predominant. As a first step in exploring untapped yeast resources of the region, this study is important for monitoring the yeast ecology in native fermentations and screening indigenous yeasts that will produce wines with regional characteristics.

Effects of Live Yeast (Saccharomyces cerevisiae Strain 1026) Supplementation on the Closure of Articular Growth Plates in Quarter Horse Foals

To avoid osteoarticular injury and to minimize farm losses in animal use and sales, breaking in young horses is ideally done as soon as growth plates have closed. An experiment was performed to determine whether dietary live yeast could shorten the time for growth plate closure. Twenty-four female Quarter Horse foals, 8 months of age, were allotted to two treatment groups (12 animals each) in a completely randomized experimental design. Horses were fed 0 or 20 g live yeast daily from 8 to 24 months of age. Diets were regular for a stud farm in the area of the study in Argentina. Foals at 8 months of age had a low baseline calcium urinary fractional excretion compatible with a primary calcium-deficient diet for this age in Argentina. Growth plate closure was evaluated by dorsum–palmar radiographs at baseline. A second evaluation was performed when foals were 12 months of age and a third evaluation when foals were 24 months of age. Blood and urine were sampled simultaneously at the same intervals to evaluate bone and mineral metabolism parameters (serum phosphorus, calcium, magnesium; alkaline phosphatase; and hydroxyproline). Calcium, magnesium, and phosphorus urinary fractional excretions were also calculated. Closure scales of distal radius growth plate data were compared using Wilcoxon test and bone markers were compared using Student t-test. Baseline radiological evaluation of growth plates showed that in 84% of foals, the distal metacarpi were either half closed or fully closed at 8 months; therefore, subsequent evaluations were limited to the distal radius. After 4 months of supplementation with live yeast at 12 months of age, a positive but nonsignificant elevation in calcium urinary fractional excretion was observed. At 24 months of age, radiographs showed a nonsignificant but positive effect in the live yeast group, with more than 50% of foals having closed distal radius growth plates. Although bone markers at 24 months of age did not differ (P > .05) between groups, some individual horses showed a positive effect with yeast supplementation.

Seashore mallow (Kosteletzkya pentacarpos) as a salt-tolerant feedstock for production of biodiesel and ethanol

Seashore mallow (Kosteletzkya pentacarpos) is a non-invasive perennial nonclonal halophytic oilseed-producing dicot that was investigated as a feedstock for production of biodiesel from seeds and ethanol from residual stem biomass. Seashore mallow seeds contained 19.3 mass % oil, which after extraction with hexane and pretreatment with catalytic sulfuric acid was converted into methyl esters in 94 mass % yield utilizing homogenous base catalysis. The principal components identified were methyl linoleate (48.9%), palmitate (24.4%) and oleate (18.3%). Fuel properties were characterized and compared to biodiesel standards ASTM D6751 and EN 14214. Also investigated were blends with petrodiesel. Lastly, seashore mallow stems were rich in neutral carbohydrates (51.8 mass %). After simultaneous saccharification and fermentation employing a native Saccharomyces cerevisiae yeast strain, the stems provided ethanol and xylose yields of 104 g/kg and 47.8 g/kg, respectively. Of the four pretreatment methodologies explored, dilute ammonium hydroxide provided the highest yield of sugars.

Knocking out ACR2 does not affect arsenic redox status in Arabidopsis thaliana: implications for as detoxification and accumulation in plants.

Many plant species are able to reduce arsenate to arsenite efficiently, which is an important step allowing detoxification of As through either efflux of arsenite or complexation with thiol compounds. It has been suggested that this reduction is catalyzed by ACR2, a plant homologue of the yeast arsenate reductase ScACR2. Silencing of AtACR2 was reported to result in As hyperaccumulation in the shoots of Arabidopsis thaliana. However, no information of the in vivo As speciation has been reported. Here, we investigated the effect of AtACR2 knockout or overexpression on As speciation, arsenite efflux from roots and As accumulation in shoots. T-DNA insertion lines, overexpression lines and wild-type (WT) plants were exposed to different concentrations of arsenate for different periods, and As speciation in plants and arsenite efflux were determined using HPLC-ICP-MS. There were no significant differences in As speciation between different lines, with arsenite accounting for >90% of the total extractable As in both roots and shoots. Arsenite efflux to the external medium represented on average 77% of the arsenate taken up during 6 h exposure, but there were no significant differences between WT and mutants or overexpression lines. Accumulation of As in the shoots was also unaffected by AtACR2 knockout or overexpression. Additionally, after exposure to arsenate, the yeast (Saccharomyces cerevisiae) strain with ScACR2 deleted showed similar As speciation as the WT with arsenite-thiol complexes being the predominant species. Our results suggest the existence of multiple pathways of arsenate reduction in plants and yeast.

Intergeneric yeast fusants with efficient ethanol production from cheese whey powder solution: Construction of aKluyveromyces marxianusandSaccharomyces cerevisiaeAY-5 hybrid

Due to its high content of lactose and abundant availability, cheese whey powder (CWP) has received much attention for ethanol production in fermentation processes. However, lactose-fermenting yeast strains including Kluyveromyces marxianus can only produce alcohol at a relatively low level, while the most commonly used distiller yeast strain Saccharomyces cerevisiae cannot ferment lactose since it lacks both β-galactosidase and the lactose permease system. To combine the unique aspects of these two yeast strains, hybrids of K. marxianus TY-22 and S. cerevisiae AY-5 were constructed by protoplast fusion. The fusants were screened and characterized by DNA content, β-galactosidase activity, ethanol tolerance, and ethanol productivity. Among the genetically stable fusants, the DNA content of strain R-1 was 6.94%, close to the sum of the DNA contents of TY-22 (3.99%) and AY-5 (3.51%). The results obtained by random-amplified polymorphic DNA analysis suggested that R-1 was a fusant between AY-5 and TY-22. During the fermentation process with CWP, the hybrid strain R-1 produced 3.8% v/v ethanol in 72 h, while the parental strain TY-22 only produced 3.1% v/v ethanol in 84 h under the same conditions.