Yeast Candida Utilis Research Articles

Interaction of a mixed yeast culture in an “autotroph-heterotroph” system with a closed atmosphere cycle and spatially separated components

The study considers an experimental model of the “autotroph-heterotroph” system with a closed atmosphere cycle, in which the heterotrophic link is a mixed yeast population. The autotrophic link is represented by the algae Chlorella vulgaris and the heterotrophic link by the yeasts Candida utilis and Candida guilliermondii. The controls are populations of Chlorella and the same yeasts isolated from the atmosphere. It has been shown that the outcome of competition in the heterotrophic link depends on the strategy of the yeast population towards the substrate and oxygen. The C. utilis population quickly utilizes the substrate as it is an r-strategist and is less sensitive to oxygen deficiency. The C. guilliermondii population consumes low concentrations of the substrate because it is a K-strategist, but it is more sensitive to oxygen deficiency. That is why, in the “autotroph-heterotroph” system with a closed gas cycle, after a considerable amount of the substrate has been consumed, the C. guilliermondii population becomes more competitive that the C. utilis population. In the culture of yeasts, isolated from the atmosphere, the C. utilis population finds itself in more favorable conditions due to oxygen deficiency. The system with a complex heterotrophic component survive longer than a system whose heterotrophic component is represented by only one yeast species. This is explained for by the positive metabolite interaction of yeasts and a more complete utilization of the substrate by a mixed culture of yeasts featuring different strategies towards the substrate.

Utilization of torula yeast (Candida utilis ) as a protein source in diets for tilapia (Oreochromis mossambicus Peters) fry

A 63 day–1 feeding trial was conducted under laboratory conditions to evaluate the effects of substituting animal protein with a mixture of plant feedstuffs including 25, 30, 35, 40 and 45% of the protein with torula yeast (Candida utilis), 20% with soybean meal and 15% with Alfalfa Leaf Protein Concentrate (ALC), in diets for tilapia (Oreochromis mossambicus Peters) fry. Feeding efficiency was compared against a diet with fish meal as the sole protein source. Diet nutritional quality was very similar independent of composition, with no differences in growth parameters, but fish fed with 30% yeast diet showed the best growth performance. Diet composition did not affect feed or protein utilization, with the best feed conversion ratio, protein efficiency ratio and apparent nitrogen utilization in the 25% yeast diet. Protein digestibility was above 80% for all diets, and no differences in carcass composition were observed. The best incidence cost was obtained with 25% yeast and the highest profit index with 30% yeast, but no statistical differences were observed with the other treatments. The results suggest that it is possible to replace up to 65% of animal protein with a mixture of plant proteins, including 30% from torula yeast, in tilapia fry diets without adverse effects on fish performance and culture profit.

Feasibility study of a compact process for biological treatment of highly soluble VOCs polluted gaseous effluent.

Volatile organic compounds (VOCs), representing a wide range of products mainly generated by industrial activity, are involved in air pollution. This study deals with a new biological treatment process of gaseous effluent combining a gas/liquid contactor called an "aero-ejector" and a membrane bioreactor. Combining these two innovative technologies enables a high elimination efficiency to be reached. We first focus on transfer phenomena characterization in a pilot installation on a laboratory scale, using a gaseous effluent polluted with a low ethanol concentration (7.1 x 10(-3) kg.m(-3)). These experiments demonstrated the good transfer performances since 90% of the ethanol was absorbed in the liquid phase in one step. After this physical characterization, the biological aspect of the system was studied using the yeast Candida utilis as microorganism. During the experiment, no ethanol was measured in the fermentation broth nor in the outlet gas, confirming the efficiency of ethanol elimination by C. utilis. The experimental procedure emerging from the present study strongly validates the suitability of this process for ethanol removal from air.

The novel non-glycosylated invertase from Candida utilis (the properties and the conditions of production and purification)

The Candida utilis yeast, which is cultivated in liquid media enriched with saccharose, synthesizes the well-known invertase of 300 kDa (EC 3.2.1.26). This enzyme is present both intracellularly in the periplasmic space and extracellularly in the culture broth. However, it was determined that the same C. utilis strain cultured in certain conditions is simultaneously capable of producing another, still unknown form of invertase with a molecular mass of 60 kDa. The presence of the latter enzymatic form was detected in cells as well as in the liquid culture medium. Both invertase forms were purified using a three-step process (ion-exchange chromatography, affinity chromatography, and preparative column electrophoresis) and named, due to their different migration ratio in polyacrylamide gel electrophoresis, F-form ( F ast; 60 kDa) and S-form ( S low; 300 kDa). The F-form of invertase was found to be nonglycosylated as opposed to the well-known S-form of invertase from the same source. The physicochemical properties of the F-form of invertase (isoelectric point, substrate specificity, pH, and temperature optima) were determined and compared with those of the S-form of the enzyme.

Oxygen requirements of the food spoilage yeast Zygosaccharomyces bailii in synthetic and complex media.

Most yeast species can ferment sugars to ethanol, but only a few can grow in the complete absence of oxygen. Oxygen availability might, therefore, be a key parameter in spoilage of food caused by fermentative yeasts. In this study, the oxygen requirement and regulation of alcoholic fermentation were studied in batch cultures of the spoilage yeast Zygosaccharomyces bailii at a constant pH, pH 3.0. In aerobic, glucose-grown cultures, Z. bailii exhibited aerobic alcoholic fermentation similar to that of Saccharomyces cerevisiae and other Crabtree-positive yeasts. In anaerobic fermentor cultures grown on a synthetic medium supplemented with glucose, Tween 80, and ergosterol, S. cerevisiae exhibited rapid exponential growth. Growth of Z. bailii under these conditions was extremely slow and linear. These linear growth kinetics indicate that cell proliferation of Z. bailii in the anaerobic fermentors was limited by a constant, low rate of oxygen leakage into the system. Similar results were obtained with the facultatively fermentative yeast Candida utilis. When the same experimental setup was used for anaerobic cultivation, in complex YPD medium, Z. bailii exhibited exponential growth and vigorous fermentation, indicating that a nutritional requirement for anaerobic growth was met by complex-medium components. Our results demonstrate that restriction of oxygen entry into foods and beverages, which are rich in nutrients, is not a promising strategy for preventing growth and gas formation by Z. bailii. In contrast to the growth of Z. bailii, anaerobic growth of S. cerevisiae on complex YPD medium was much slower than growth in synthetic medium, which probably reflected the superior tolerance of the former yeast to organic acids at low pH.

Interrelations of the yeast Candida utilis and Cr(VI): metal reduction and its distribution in the cell and medium

An effect of chromium(VI) ions on the growth and bioaccumulation properties of growing cells of Candida Utilis was studied. Molasses media for yeast growth containing 20 g glucose l −1 and 50+500 mg Cr(VI) l −1 were used in batch cultivation. Addition of 100 mg Cr(VI) l −1 resulted in a threefold decrease in the cell concentration, as compared with the culture grown without metal. Cr(VI) inhibited culture growth in a concentration-dependent manner, this dependence was not linear. Glucose consumption by growing cells depended on the initial Cr(VI) concentration in the medium and correlated with growth activity. No inhibitory effect of high Cr(VI) concentrations on the activity of some exo-enzymes of C. utilis cells was observed. During C. utilis cultivation, Cr(VI) was partially reduced to an ultimate Cr(III) in all tested ranges of the Cr(VI) concentrations. The specific chromium uptake by cells was detected in biomass that grew in the presence of 100+300 mg Cr(VI) l −1 in a period of 48–96 h. The highest values were achieved after 96 h growth in the presence of 200 and 300 mg Cr(VI) l −1, and were 7.3 and 7.2 mg Cr g dw l −1, respectively. Electron microscopic observations showed morphological changes in yeast cells to be more pronounced upon culture growth with 100–300 mg Cr(VI) l −1 than in the cells subjected to higher metal concentrations. The conclusion has been made that the mechanism of the Cr(VI) toxic effect on the growing yeast cells can vary in dependence of the metal concentration.

Isolation and sequence of the MIG1 homologue from the yeast Candida utilis.

The Mig1p repressor from the food yeast Candida utilis has been isolated using a homologous PCR hybridization probe. This probe was amplified with two sets of degenerate primers designed on the basis of highly conserved motifs in the DNA-binding region (zinc-finger domain) from yeast Mig1p and fungi CreA repressors. The cloned gene was sequenced and found to encode a polypeptide of 345 amino acids which shows significant identity with other yeast and fungus repressors in the DNA-binding domain and also with the yeast Mig1 proteins in the C-terminal region (effector domain). The MIG1 repressor gene from C. utilis was able to complement functionally the mig1 mutation of S. cerevisiae. The sequence presented here has been deposited in the EMBL data library under Accession No. AJ277830.

Differential uptake of fumarate by Candida utilis and Schizosaccharomyces pombe.

The dicarboxylic acid fumarate is an important intermediate in cellular processes and also serves as a precursor for the commercial production of fine chemicals such as L-malate. Yeast species differ remarkably in their ability to degrade extracellular dicarboxylic acids and to utilise them as their only source of carbon. In this study we have shown that the yeast Candida utilis effectively degraded extracellular fumarate and L-malate, but glucose or other assimilable carbon sources repressed the transport and degradation of these dicarboxylic acids. The transport of both dicarboxylic acids was shown to be strongly inducible by either fumarate or L-malate while kinetic studies suggest that the two dicarboxylic acids are transported by the same transporter protein. In contrast, Schizosaccharomyces pombe effectively degraded extracellular L-malate, but not fumarate, in the presence of glucose or other assimilable carbon sources. The Sch. pombe malate transporter was unable to transport fumarate, although fumarate inhibited the uptake of L-malate.

The influence of Debaryomyces hansenii and Candida utilis on the aroma formation in garlic spiced fermented sausages and model minces

The influence of the yeast starter cultures Debaryomyces hansenii and Candida utilis on fermented meat aroma was studied in model minces and in commercial-type fermented sausages. Volatile compounds from model minces and sausages were collected using diffusive and dynamic headspace sampling respectively and were identified by gas chromatography/mass spectrometry (GC/MS). A triangle test was carried out on the sausages to detect whether the yeast influenced the sausage odour. C. utilis demonstrated high metabolic activity in the model minces, producing several volatile compounds, in particularly esters. C. utilis also seemed to ferment the amino acids valine, isoleucine and leucine into compounds important for the aroma of sausages. D. hansenii on the contrary, had very little effect on the production of volatile compounds in the model minces. In the sausage experiment both yeast cultures died out before the ripening process ended and the sensory analysis showed only a slight difference between the sausages. A fungistatic test of the garlic powder added to the sausages indicated that garlic inhibits the growth of the yeast starter cultures.

A novel hemiacetal dehydrogenase activity involved in ethyl acetate synthesis in Candida utilis

Acetate ester synthesis was studied in vitro with the ethyl acetate-producing yeast Candida utilis. The level of enzyme activity observed for the NAD +-dependent hemiacetal dehydrogenase acting on hemiacetal, which was produced non-enzymatically from an alcohol and an aldehyde, was much greater than that for the other enzyme involved in ester synthesis, alcohol acetyltransferase. The level of ethyl acetate synthesis in vivo approximately paralleled the hemiacetal dehydrogenase (HADH) activity. The results suggest that the main pathway for ethyl acetate synthesis in C. utilis involves a novel hemiacetal dehydrogenase activity.

Continuous cultivation of the yeast Candida utilis at different dilution rates on pineapple cannery effluent

Candida utilis was grown on a pineapple cannery effluent in a chemostat at dilution rates ranging between 0.05 and 0.65 h−1 to establish optimal conditions for biomass production and chemical oxygen demand (COD) reduction. Sucrose, fructose and glucose were the main sugars in the effluent. Maximum value for cell yield coefficient and productivity were (0.686, gx/gs) and (2.96, gx/l/h) at a dilution rate of 0.425 and 0.475 h−1, respectively, while maximum COD reduction (98%) was attained at a dilution rate of 0.1 h−1. The maintenance coefficient attained a value of (0.093, gs/gx/h). An increase in dilution rate produced a higher protein content of the biomass.

PREPARATION OF AN IMITATION SOY SAUCE FROM HYDROLYZED DRIED YEAST CANDIDA UTILIS

Acid hydrolysis of dried fodder's yeast Candida utilis was studied by a central composite design taking into account three independent variables: temperature (105-165C), processing time (30-70 min) and HCI concentration (7.5-17.5%). Response variables were: color, dry matter solubilization and amino nitrogen. Temperature was the most significant variable followed by acid concentration and processing time. Response surfaces were generated from fitted polynomials. Higher temperatures to prevent amino nitrogen degradation were not investigated. As different effects were found for color development and amino nitrogen, a combined function was tested which yields the behavior of all responses simultaneously. The best conditions were: 150C, 15% HCl and 40 min. After neutralization with 6N NaOH and centrifugation, the raw sauce was supplemented with 12% NaCl, 0.1% monosodium glutamate, 2% vinegar and 0.1% sodium benzoate and evaluated by sensory analysis for eight distinct attributes. Imitation sauce from yeast was at least as good as commercial hydrolyzed soybean sauce.

Cloning and sequence analysis of the gene encoding invertase (INV1) from the yeast Candida utilis.

The gene INV1 encoding invertase from the yeast Candida utilis has been cloned using a homologous PCR hybridization probe, amplified with two sets of degenerate primers designed considering sequence comparisons between yeast invertases. The cloned gene was sequenced and found to encode a polypeptide of 533 amino acids that contain a 26 amino-acid signal peptide and 12 potential N-glycosylation sites. The nucleotide sequences of the 5' and 3' non-coding regions were found to contain motifs probably involved in initiation, regulation and termination of gene transcription. The amino-acid sequence shows significant identity with other yeast, bacterial and plant beta-fructofuranosidases. The INV1 gene from C. utilis was able to complement functionally the suc2 mutation of S. cerevisiae.

Cloning and sequence analysis of the gene encoding invertase (INV1) from the yeast Candida utilis

Cloning and sequence analysis of the gene encoding invertase (INV1) from the yeast Candida utilis

Development of an integrative DNA transformation system for the yeast Candida utilis

We report here the development of an auxotrophic transformation system for the food yeast Candida utilis. To facilitate molecular studies in Candida utilis, we isolated auxotrophic strains for uracil biosynthesis by the combination of NTG-mutagenesis and 5-fluorotic acid (FOA) selection. The ura − mutation could be functionally complemented by the homologous URA3 gene. We used both, LiAc and electroporation methods to direct insertions at the ura3 locus through homologous recombination.

Development of an integrative DNA transformation system for the yeast Candida utilis.

We report here the development of an auxotrophic transformation system for the food yeast Candida utilis. To facilitate molecular studies in Candida utilis, we isolated auxotrophic strains for uracil biosynthesis by the combination of NTG-mutagenesis and 5-fluorotic acid (FOA) selection. The ura-mutation could be functionally complemented by the homologous URA3 gene. We used both, LiAc and electroporation methods to direct insertions at the ura3 locus through homologous recombination.

Metabolic engineering for the production of carotenoids in non-carotenogenic bacteria and yeasts

The crt gene clusters responsible for the biosynthesis of carotenoids such as lycopene, β-carotene and astaxanthin have been isolated from carotenogenic bacteria such as Erwinia species and the marine bacterium Agrobacterium aurantiacum. The functions of the individual genes have been identified. The first substrate of the enzymes encoded by the Erwinia crt clusters is farnesyl pyrophosphate which is not only the precursor for carotenoid biosynthesis but also sterols, dolichols and other numerous isoprenoid compounds. Escherichia coli does not naturally synthesize carotenoids, but by using the carotenogenic genes recombinant strains accumulating lycopene, β-carotene and astaxanthin have been produced. Other non-carotenogenic bacteria such as Zymomonas mobilis have also been engineered to produce β-carotene by the introduction of the corresponding crt genes. A gene capable of enhancing carotenoid levels in E. coli has also been isolated from cDNA libraries of the yeast Phaffia rhodozyma and the green alga Haematococcus pluvialis. This gene has been found to encode an isopentenyl pyrophophate isomerase. It has further been shown that the edible yeasts Candida utilis as well as Saccharomyces cerevisiae, which possess no carotenoid biosynthetic pathway, acquire the ability to produce carotenoids, when the carotenogenic genes are expressed under the control of yeast-derived promoters and terminators. It has been observed in the yeasts S. cerevisiae and C. utilis carrying the lycopene biosynthesis genes that ergosterol content is decreased by 10 and 35%, respectively. It is therefore likely that the carbon flux for the ergosterol biosynthesis has been partially directed from farnesyl pyrophosphate to a new pathway for the lycopene biosynthesis. Further, the expression of a truncated gene which codes for the catalytic domain of the endogenous 3-hydroxy-3-methylglutaryl coenzyme A reductase, has been found to be effective for enhancing carotenoid levels in the yeast C. utilis.

High-level expression of a sweet protein, monellin, in the food yeast Candida utilis.

We describe the heterologous expression of monellin in the yeast Candida utilis. A single-chain monellin gene was expressed under the control of the glyceraldehyde-3-phosphate dehydrogenase [correction of decarboxylase] gene promoter from C. utilis. A promoter-deficient marker gene allowed high-copy-number integration of vectors into either the rDNA locus or the URA3 gene locus. Monellin was produced at a high level, accounting for > 50% of the soluble protein. No significant decrease in the production level of monellin was detected in transformants after 50 generations of nonselective growth.

Purification and characterization of an invertase from Candida utilis: comparison with natural and recombinant yeast invertases

A periplasmic invertase from the yeast Candida utilis was purified to homogeneity from cells fully derepressed for invertase synthesis. The enzyme was purified by successive Sephacryl S-300, and affinity chromatography and shown to be a dimeric glycoprotein composed of two identical monomer subunits with an apparent molecular mass of 150 kDa. After EndoH treatment, the deglycosylated protein showed an apparent molecular weight of 60 kDa. The apparent K m values for sucrose and raffinose were 11 and 150 mM, respectively, similar to those reported in Saccharomyces cerevisiae. The range of optimum temperature was 60–75°C. The optimum pH was 5.5 and the enzyme was stable over pH range 3–6.

Mechanism of glucose and maltose transport in plasma-membrane vesicles from the yeast Candida utilis.

Transport of glucose and maltose was studied in plasma-membrane vesicles from Candida utilis. The yeast was grown on a mixture of glucose and maltose in aerobic carbon-limited continuous cultures which enabled transport to be studied for both sugars with the same vesicles. Vesicles were prepared by fusion of isolated plasma membranes with proteoliposomes containing bovine heart cytochrome c oxidase as a proton-motive-force-generating system. Addition of reduced cytochrome c generated a proton-motive force, consisting of a membrane potential, negative inside, and a pH gradient, alkaline inside. Energization led to accumulation of glucose and maltose in these vesicles, reaching accumulation ratios of about 40-50. Accumulation also occurred in the presence of valinomycin or nigericin, but was prevented by a combination of the two ionophores or by uncoupler, showing that glucose and maltose transport are dependent on the proton-motive force. Comparison of sugar accumulation with quantitative data on the proton-motive force indicated a 1:1 H+/sugar stoichiometry for both transport systems. Efflux of accumulated glucose was observed on dissipation of the proton-motive force. Exchange and counterflow experiments confirmed the reversible character of the H+-glucose symporter. In contrast, uncoupler or a mixture of valinomycin plus nigericin induced only a slow efflux of accumulated maltose. Moreover under counterflow conditions, the expected transient accumulation was small. Thus the H+-maltose symporter has some characteristics of a carrier that is not readily reversible. It is concluded that in C. utilis the transport systems for glucose and maltose are both driven by the proton-motive force, but the mechanisms are different.