Relationship between pellet formation by Aspergillus oryzae strain KB and the production of ß-fructofuranosidase with high transfructosylation activity.

Aspergillus oryzae KB produces two ß-fructofuranosidases (F1 and F2). F1 has high transfructosylation activity (Ut) to produce fructooligosaccharides. F2 has high hydrolysis activity (Uh), releasing glucose and fructose. It is desirable to selectively produce F1, which can be used for production of fructooligosaccharides. Here, the relationship between filamentous pellet size and selective production of F1 in liquid culture was investigated. Our finding revealed that: (i) The mean particle size of pellets (5.88 ± 1.36 mm) was larger, and the ratio of Ut to Uh was improved (Ut/Uh = 5.0) in 10% sucrose medium compared with 1% sucrose medium (pellet size = 2.60 ± 0.37 mm; Ut/Uh = 0.96). (ii) The final culture pH of the 1% sucrose medium was 8.7; on controlling the pH of 1% sucrose medium at 5.0, increased pellet size (9.69 ± 2.01 mm) and Ut/Uh (7.8) were observed. (iii) When 3% glycerin was used as carbon source, the pellet size decreased to 1.09 ± 0.33 mm and Ut/Uh was 0.57. (iv) In medium containing 1% sucrose, the pellet size was dependent on the number of spores used in the culture inoculum, but, in these experiments, Ut/Uh was almost constant (1.05 ± 0.08). Collectively, the data show that the value of Ut/Uh is proportional to the pellet size when liquid culture of A. oryzae strain KB is performed in some conditions (such as in the presence of high sucrose concentration, low pH, or added Tween surfactant), but in other conditions Ut/Uh is independent of pellet size.

Overproduction of a ß-fructofuranosidase1 with a high FOS synthesis activity for efficient biosynthesis of fructooligosaccharides.

Aureobasidium melanogenum 11-1 was found to be able to produce over 281.7 ±â€¯7.1 U/mL of ß-fructofuranosidase activity. The protein deduced from the cloned ß-fructofuranosidase1 gene had the conserved motif A (IGDP), motif D (RDP) and motif E (ET) and 11 N-glycosylation sites, indicating it was a ß-fructofuranosidase with the high-level fructooligosaccharide (FOS) biosynthesis. Overexpression of the ß-fructofuranosidase1 gene in the yeast strain 11-1 made a tranformant 33 produce 557.7 U/mL of ß-fructofuranosidase activity. The molecular weight of the ß-fructofuranosidase1 in which all the carbohydrates were removed by the Endo-H was 82.4 kDa. Within 7 h of the transfructosylation reaction, the yield of FOS was 0.66 g of FOS/g of sucrose and percentages of GF2, GF3 and GF4 were 79.5%, 18.9% and 1.6%. This demonstrated that the ß-fructofuranosidase1 and the transformant 33 had highly potential applications in biotechnology for FOS production.

Biochemical characterization and evaluation of invertases produced from Saccharomyces cerevisiae CAT-1 and Rhodotorula mucilaginosa for the production of fructooligosaccharides.

Invertases are used for several purposes; one among these is the production of fructooligosaccharides. The aim of this study was to biochemically characterize invertase from industrial Saccharomyces cerevisiae CAT-1 and Rhodotorula mucilaginosa isolated from Cerrado soil. The optimum pH and temperature were 4.0 and 70 °C for Rhodotorula mucilaginosa invertase and 4.5 and 50 °C for Saccharomyces cerevisiae invertase. The pH and thermal stability from 3.0 to 10.5 and 75 °C for R. mucilaginosa invertase, respectively. The pH and thermal stability for S. cerevisiae CAT-1 invertase from 3.0 to 7.0, and 50 °C, respectively. Both enzymes showed good catalytic activity with 10% of ethanol in reaction mixture. The hydrolysis by invertases occurs predominantly when sucrose concentrations are ≤5%. On the other hand, the increase in the concentration of sucrose to levels above 10% results in the highest transferase activity, reaching about 13.3 g/L of nystose by S. cerevisiae invertase and 12.6 g/L by R. mucilaginosa invertase. The results demonstrate the high structural stability of the enzyme produced by R. mucilaginosa, which is an extremely interesting feature that would enable the application of this enzyme in industrial processes.

Rapid evaluation of 1-kestose producing ß-fructofuranosidases from Aspergillus species and enhancement of 1-kestose production using a PgsA surface-display system.

1-Kestose is a key prebiotic fructooligosaccharide (FOS) sugar. Some ß-fructofuranosidases (FFases) have high transfructosylation activity, which is useful for manufacturing FOS. Therefore, obtaining FFases that produce 1-kestose efficiently is important. Here, we established a rapid FFase evaluation method using Escherichia coli that display different FFases fused to a PgsA anchor protein from Bacillus subtilis. E. coli cell suspensions expressing the PgsA-FFase fusion efficiently produce FOS from sucrose. Using this screening technique, we found that the E. coli transformant expressing Aspergillus kawachii FFase (AkFFase) produced a larger amount of 1-kestose than those expressing FFases from A. oryzae and A. terreus. Saturation mutagenesis of AkFFase was performed, and the mutant G85W was obtained. The E. coli transformant expressing AkFFase G85W markedly increased production of 1-kestose. Our results indicate that the surface display technique using PgsA is useful for screening of FFases, and AkFFase G85W is likely to be suitable for 1-kestose production. ABBREVIATIONS: AkFFase: Aspergillus kawachii FFase; AoFFase: Aspergillus oryzae FFase; AtFFase: Aspergillus terreus FFase; FFase: ß-fructofuranosidase; FOS: fructooligosaccharide; fructosylnystose: 1F-ß-fructofuranosylnystose.

Synthesis of fructooligosaccharides (FosA) and inulin (InuO) by GH68 fructosyltransferases from Bacillus agaradhaerens strain WDG185.

Fructooligosaccharides (FOS) and inulin, composed of ß-2-1 linked fructose units, have a broad range of industrial applications. They are known to have various beneficial health effects and therefore have broad application potential in nutrition. For (modified) inulin also for non-food purposes more applications are arising. Examples are carboxymethylated inulin as anti-scalant and carboymlated inulin as emulsifiers. Various plants synthesize FOS and/or inulin type of fructans. However, isolating of FOS and inulin from plants is challenging due to for instance varying chains length. There is an increasing demand for FOS and inulin oligosaccharides and alternative procedures for their synthesis are attractive. We identified and characterized two fructosyltransferases from Bacillus agaradhaerens WDG185. FosA, a ß-fructofuranosidase, synthesises short chain fructooligosaccharides (GF2-GF4) at high sucrose concentration, whereas InuO, an inulosucrase, synthesises a broad range of inulooligosaccharides (GF2-GF24) from sucrose, very similar to plant derived inulin. FosA and InuO showed activity over a broad pH range from 6 to 10 and optimal temperature at 60°C. Calcium ions and EDTA were found to have no effect on the activity of both enzymes. Kinetic analysis showed that only at relatively low substrate concentrations both enzymes showed Michaelis-Menten type of kinetics for total and transglycosylation activity. Both enzymes showed increased transglycosylation upon increasing substrate concentrations. These are the first examples of the molecular and biochemical characterization of a ß-fructofuranosidase (FosA) and an inulosucrase enzyme (InuO) and its product from a Bacillus agaradhaerens strain.

Bacterial invertases: Occurrence, production, biochemical characterization, and significance of transfructosylation.

Invertase or ß-D-fructofuranoside fructohydrolase (EC 3.2.1.26) was one of the foremost enzyme biocatalysts and established the primary concepts of most enzyme-kinetic principles. Invertases are glycoside hydrolases and occur mostly in microorganisms. Among microbial strains, for many decades yeast species have been extensively researched for invertase production, characterization, and applications in industries. Besides, limited literature is available on invertases from bacterial strains. The enzymic and molecular biological reports from bacterial invertases are scarce. In this minireview, occurrence, production, biochemical properties, and significance of transfructosylation of bacterial invertases are reported.

Optimizing culture conditions for production of intra and extracellular inulinase and invertase from Aspergillus niger ATCC 20611 by response surface methodology (RSM)

Abstract The aim of this study was obtain a model that maximizes growth and production of inulinase and invertase by Aspergillus niger ATCC 20611, employing response surface methodology (RSM). The RSM with a five-variable and three-level central composite design (CCD) was employed to optimize the medium composition. Results showed that the experimental data could be appropriately fitted into a second-order polynomial model with a coefficient of determination (R2) more than 0.90 for all responses. This model adequately explained the data variation and represented the actual relationships between the parameters and responses. The pH and temperature value of the cultivation medium were the most significant variables and the effects of inoculum size and agitation speed were slightly lower. The intra-extracellular inulinase, invertase production and biomass content increased 10-32 fold in the optimized medium condition (pH 6.5, temperature 30 °C, 6% (v/v), inoculum size and 150 rpm agitation speed) by RSM compared with medium optimized through the one-factor-at-a-time method. The process development and intensification for simultaneous production of intra-extracellular inulinase (exo and endo inulinase) and invertase from A. niger could be used for industrial applications.

Optimizing culture conditions for production of intra and extracellular inulinase and invertase from Aspergillus niger ATCC 20611 by response surface methodology (RSM).

The aim of this study was obtain a model that maximizes growth and production of inulinase and invertase by Aspergillus niger ATCC 20611, employing response surface methodology (RSM). The RSM with a five-variable and three-level central composite design (CCD) was employed to optimize the medium composition. Results showed that the experimental data could be appropriately fitted into a second-order polynomial model with a coefficient of determination (R2) more than 0.90 for all responses. This model adequately explained the data variation and represented the actual relationships between the parameters and responses. The pH and temperature value of the cultivation medium were the most significant variables and the effects of inoculum size and agitation speed were slightly lower. The intra-extracellular inulinase, invertase production and biomass content increased 10-32 fold in the optimized medium condition (pH 6.5, temperature 30°C, 6% (v/v), inoculum size and 150rpm agitation speed) by RSM compared with medium optimized through the one-factor-at-a-time method. The process development and intensification for simultaneous production of intra-extracellular inulinase (exo and endo inulinase) and invertase from A. niger could be used for industrial applications.

Effects of nonionic surfactants on pellet formation and the production of ß-fructofuranosidases from Aspergillus oryzae KB.

Aspergillus oryzae KB produces two ß-fructofuranosidases (F1 and F2). F1 has high transferring activity and produces fructooligosaccharides from sucrose. Mycelial growth pellets were altered by the addition of Tween 20, 40 and 80 (HLB=16.7, 15.6 and 15.0, respectively) in liquid medium cultures to form small spherical pellets. The particle size of the pellets decreased with the HLB value, which corresponds to an increase in surfactant hydrophobicity. Selective F1 production and pellet size were maximized using Tween 20. Adding polyoxyethylene oleyl ethers (POEs) with various degrees of polymerization (2, 7, 10, 20 and 50: HLB=7.7, 10.7, 14.7, 17.2 and 18.2, respectively) was investigated. A minimum mean particle size was obtained using a POE with DP=10, HLB=14.7. The POE surfactants had little effect on the selective production of F1. The formation of filamentous pellets depended on the surfactant HLB value, and F1 enzymes were produced most efficiently using Tween 20.

Cytokinins and Expression of SWEET, SUT, CWINV and AAP Genes Increase as Pea Seeds Germinate.

Transporter genes and cytokinins are key targets for crop improvement. These genes are active during the development of the seed and its establishment as a strong sink. However, during germination, the seed transitions to being a source for the developing root and shoot. To determine if the sucrose transporter (SUT), amino acid permease (AAP), Sugar Will Eventually be Exported Transporter (SWEET), cell wall invertase (CWINV), cytokinin biosynthesis (IPT), activation (LOG) and degradation (CKX) gene family members are involved in both the sink and source activities of seeds, we used RT-qPCR to determine the expression of multiple gene family members, and LC-MS/MS to ascertain endogenous cytokinin levels in germinating Pisum sativum L. We show that genes that are actively expressed when the seed is a strong sink during its development, are also expressed when the seed is in the reverse role of being an active source during germination and early seedling growth. Cytokinins were detected in the imbibing seeds and were actively biosynthesised during germination. We conclude that, when the above gene family members are targeted for seed yield improvement, a downstream effect on subsequent seed germination or seedling vigour must be taken into consideration.

Double mutation of Saccharomyces cerevisiae for enhanced β-D-fructofuranosidase fructohydrolase productivity and application of growth kinetics for parametric significance analysis

Abstract The kinetics of an extracellular β-D-fructofuranosidase fructohydrolase production by Saccharomyces cerevisiae in a chemically defined medium, i.e., sucrose peptone agar yeast extract at pH 6, was investigated. The wild-type was treated with a chemical mutagen, methyl methane sulfonate. Among the six mutants isolated, methyl methane sulfonate-V was found to be a better enzyme producing strain (52 ± 2.4a U/mL). The maximum production (74 ± 3.1a U/mL) was accomplished after at 48 h (68 ± 2.7a mg/mL protein). The mutants were stabilized at low levels of 5-fluoro-cytocine and the viable ones were further processed for optimization of cultural conditions and nutritional requirements. The sucrose concentration, incubation period and pH were optimized to be 30 g/L, 28 °C, and 6.5, respectively. The methyl methane sulfonate-V exhibited an improvement of over 10 folds in enzyme production when 5 g/L ammonium sulfate was used as a nitrogen source. Thin layer chromatography and high-performance liquid chromatography analysis illustrated the optimal enzyme activity supported by the higher rate of hydrolysis of sucrose into monosaccharides, particularly α-D-glucose and β-D-fructose. The values for Qp (2 ± 0.12c U/mL/h) and Yp/s (4 ± 1.24b U/g) of the mutant were considerably increased in comparison with other yeast strains (both isolates and viable mutants). The mutant could be exploited for enzyme production over a wider temperature range (26–34 °C), with significantly high enzyme activity (LSD 0.048, HS) at the optimal temperature.

Double mutation of Saccharomyces cerevisiae for enhanced ß-d-fructofuranosidase fructohydrolase productivity and application of growth kinetics for parametric significance analysis.

The kinetics of an extracellular ß-d-fructofuranosidase fructohydrolase production by Saccharomyces cerevisiae in a chemically defined medium, i.e., sucrose peptone agar yeast extract at pH 6, was investigated. The wild-type was treated with a chemical mutagen, methyl methane sulfonate. Among the six mutants isolated, methyl methane sulfonate-V was found to be a better enzyme producing strain (52±2.4(a)U/mL). The maximum production (74±3.1(a)U/mL) was accomplished after at 48h (68±2.7(a)mg/mL protein). The mutants were stabilized at low levels of 5-fluoro-cytocine and the viable ones were further processed for optimization of cultural conditions and nutritional requirements. The sucrose concentration, incubation period and pH were optimized to be 30g/L, 28°C, and 6.5, respectively. The methyl methane sulfonate-V exhibited an improvement of over 10 folds in enzyme production when 5g/L ammonium sulfate was used as a nitrogen source. Thin layer chromatography and high-performance liquid chromatography analysis illustrated the optimal enzyme activity supported by the higher rate of hydrolysis of sucrose into monosaccharides, particularly α-d-glucose and ß-d-fructose. The values for Qp (2±0.12(c)U/mL/h) and Yp/s (4±1.24(b)U/g) of the mutant were considerably increased in comparison with other yeast strains (both isolates and viable mutants). The mutant could be exploited for enzyme production over a wider temperature range (26-34°C), with significantly high enzyme activity (LSD 0.048, HS) at the optimal temperature.

Hexokinase 2 Is an Intracellular Glucose Sensor of Yeast Cells That Maintains the Structure and Activity of Mig1 Protein Repressor Complex.

Hexokinase 2 (Hxk2) fromSaccharomyces cerevisiaeis a bi-functional enzyme, being both a catalyst in the cytosol and an important regulator of the glucose repression signal in the nucleus. Despite considerable recent progress, little is known about the regulatory mechanism that controls nuclear Hxk2 association with theSUC2promoter chromatin and how this association is necessary forSUC2gene repression. Our data indicate that in theSUC2promoter context, Hxk2 functions through a variety of structurally unrelated factors, mainly the DNA-binding Mig1 and Mig2 repressors and the regulatory Snf1 and Reg1 factors. Hxk2 sustains the repressor complex architecture maintaining transcriptional repression at theSUC2gene. Using chromatin immunoprecipitation assays, we discovered that the Hxk2 in its open configuration, at low glucose conditions, leaves the repressor complex that induces its dissociation and promotesSUC2gene expression. In high glucose conditions, Hxk2 adopts a close conformation that promotes Hxk2 binding to the Mig1 protein and the reassembly of theSUC2repressor complex. Additional findings highlight the possibility that Hxk2 constitutes an intracellular glucose sensor that operates by changing its conformation in response to cytoplasmic glucose levels that regulate its interaction with Mig1 and thus its recruitment to the repressor complex of theSUC2promoter. Thus, our data indicate that Hxk2 is more intimately involved in gene regulation than previously thought.

A unique, highly conserved secretory invertase is differentially expressed by promastigote developmental forms of all species of the human pathogen, Leishmania.

Leishmania are protozoan pathogens of humans that exist as extracellular promastigotes in the gut of their sand fly vectors and as obligate intracellular amastigotes within phagolysosomes of infected macrophages. Between infectious blood meal feeds, sand flies take plant juice meals that contain sucrose and store these sugars in their crop. Such sugars are regurgitated into the sand fly anterior midgut where they impact the developing promastigote parasite population. In this report we showed that promastigotes of all Leishmania species secreted an invertase/sucrase enzyme during their growth in vitro. In contrast, neither L. donovani nor L. mexicana amastigotes possessed any detectable invertase activity. Importantly, no released/secreted invertase activity was detected in culture supernatants from either Trypanosoma brucei or Trypanosoma cruzi. Using HPLC, the L. donovani secretory invertase was isolated and subjected to amino acid sequencing. Subsequently, we used a molecular approach to identify the LdINV and LmexINV genes encoding the ~72 kDa invertases produced by these organisms. Interestingly, we identified high fidelity LdINV-like homologs in the genomes of all Leishmania sp. but none were present in either T. brucei or T. cruzi. Northern blot and RT-PCR analyses showed that these genes were developmentally/differentially expressed in promastigotes but not amastigotes of these parasites. Homologous transfection studies demonstrated that these genes in fact encoded the functional secretory invertases produced by these parasites. Cumulatively, our results suggest that these secretory enzymes play critical roles in the survival/growth/development and transmission of all Leishmania parasites within their sand fly vector hosts.

[Export of an invertase by yeast cells (Candida utilis)].

Export and accumulation of various forms of invertase (EC 3.2.1.26) in the cell wall and culture liquid of the yeast Candida utilis was investigated. It was found that the high-molecular-weight CW-form of invertase is present in the cell wall. This form is not exported into the culture liquid, and it is by a third more glycosylated than the previously described exported S-form. It was shown that one of the two liquid forms of invertase exported into the culture-the glycosylated S-form--is retained in the cell wall, while the other one--the nonglycosylated F-form--was not detected in the cell wall. Based on these results, as well as data on the distribution dynamics of the enzyme in the culture liquid and in the cell wall during different growth stages of a yeast culture, we suggested that the nonglycosylated form was exported into the culture liquid via the zone of abnormal cell wall permeability and the glycosylated forms of this enzyme (both exported and nonexported) did not use this pathway (the degree of N-glycosylation is an important factor determining the final localization of the enzyme).

Production and characterization of a novel yeast extracellular invertase activity towards improved dibenzothiophene biodesulfurization.

The main goal of this work was the production and characterization of a novel invertase activity from Zygosaccharomyces bailii strain Talf1 for further application to biodesulfurization (BDS) in order to expand the exploitable alternative carbon sources to renewable sucrose-rich feedstock. The maximum invertase activity (163 U ml(-1)) was achieved after 7 days of Z. bailii strain Talf1 cultivation at pH 5.5-6.0, 25 °C, and 150 rpm in Yeast Malt Broth with 25 % Jerusalem artichoke pulp as inducer substrate. The optimum pH and temperature for the crude enzyme activity were 5.5 and 50 °C, respectively, and moreover, high stability was observed at 30 °C for pH 5.5-6.5. The application of Talf1 crude invertase extract (1 %) to a BDS process by Gordonia alkanivorans strain 1B at 30 °C and pH 7.5 was carried out through a simultaneous saccharification and fermentation (SSF) approach in which 10 g l(-1) sucrose and 250 µM dibenzothiophene were used as sole carbon and sulfur sources, respectively. Growth and desulfurization profiles were evaluated and compared with those of BDS without invertase addition. Despite its lower stability at pH 7.5 (loss of activity within 24 h), Talf1 invertase was able to catalyze the full hydrolysis of 10 g l(-1) sucrose in culture medium into invert sugar, contributing to a faster uptake of the monosaccharides by strain 1B during BDS. In SSF approach, the desulfurizing bacterium increased its µmax from 0.035 to 0.070 h(-1) and attained a 2-hydroxybiphenyl productivity of 5.80 µM/h in about 3 days instead of 7 days, corresponding to an improvement of 2.6-fold in relation to the productivity obtained in BDS process without invertase addition.

The genetics of a putative social trait in natural populations of yeast.

The sharing of secreted invertase by yeast cells is a well-established laboratory model for cooperation, but the only evidence that such cooperation occurs in nature is that the SUC loci, which encode invertase, vary in number and functionality. Genotypes that do not produce invertase can act as 'cheats' in laboratory experiments, growing on the glucose that is released when invertase producers, or 'cooperators', digest sucrose. However, genetic variation for invertase production might instead be explained by adaptation of different populations to different local availabilities of sucrose, the substrate for invertase. Here we find that 110 wild yeast strains isolated from natural habitats, and all contained a single SUC locus and produced invertase; none were 'cheats'. The only genetic variants we found were three strains isolated instead from sucrose-rich nectar, which produced higher levels of invertase from three additional SUC loci at their subtelomeres. We argue that the pattern of SUC gene variation is better explained by local adaptation than by social conflict.

Cloning, 3D modeling and expression analysis of three vacuolar invertase genes from cassava (Manihot Esculenta Crantz).

Vacuolar invertase is one of the key enzymes in sucrose metabolism that irreversibly catalyzes the hydrolysis of sucrose to glucose and fructose in plants. In this research, three vacuolar invertase genes, named MeVINV1-3, and with 653, 660 and 639 amino acids, respectively, were cloned from cassava. The motifs of NDPNG (ß-fructosidase motif), RDP and WECVD, which are conserved and essential for catalytic activity in the vacuolar invertase family, were found in MeVINV1 and MeVINV2. Meanwhile, in MeVINV3, instead of NDPNG we found the motif NGPDG, in which the three amino acids GPD are different from those in other vacuolar invertases (DPN) that might result in MeVINV3 being an inactivated protein. The N-terminal leader sequence of MeVINVs contains a signal anchor, which is associated with the sorting of vacuolar invertase to vacuole. The overall predicted 3D structure of the MeVINVs consists of a five bladed ß-propeller module at N-terminus domain, and forms a ß-sandwich module at the C-terminus domain. The active site of the protein is situated in the ß-propeller module. MeVINVs are classified in two subfamilies, α and ß groups, in which α group members of MeVINV1 and 2 are highly expressed in reproductive organs and tuber roots (considered as sink organs), while ß group members of MeVINV3 are highly expressed in leaves (source organs). All MeVINVs are highly expressed in leaves, while only MeVINV1 and 2 are highly expressed in tubers at cassava tuber maturity stage. Thus, MeVINV1 and 2 play an important role in sucrose unloading and starch accumulation, as well in buffering the pools of sucrose, hexoses and sugar phosphates in leaves, specifically at later stages of plant development.

Kinetics and thermodynamics of ethanol production by Saccharomyces cerevisiae MLD10 using molasses.

In this study, we have used ultraviolet (UV) and γ-ray induction to get a catabolite repression resistant and thermotolerant mutant with enhanced ethanol production along with optimization of sugar concentration and temperature of fermentation. Classical mutagenesis in two consecutive cycles of UV- and γ-ray-induced mutations evolved one best catabolite-resistant and thermotolerant mutant Saccharomyces cerevisiae MLD10 which showed improved ethanol yield (0.48 ± 0.02 g g(-1)), theoretical yield (93 ± 3%), and extracellular invertase productivity (1,430 ± 50 IU l(-1) h(-1)), respectively, when fermenting 180 g sugars l(-1) in molasses medium at 43 °C in 300 m(3) working volume fermenter. Ethanol production was highly dependent on invertase production. Enthalpy (ΔH*) (32.27 kJ M(-1)) and entropy (ΔS*) (-202.88 J M(-1) K(-1)) values at 43 °C by the mutant MLD10 were significantly lower than those of ß-glucosidase production by a thermophilic mutant derivative of Thermomyces lanuginosus. These results confirmed the enhanced production of ethanol and invertase by this mutant derivative. These studies proved that mutant was significantly improved for ethanol production and was thermostable in nature. Lower fermentation time for ethanol production and maintenance of ethanol production rates (3.1 g l(-1) h(-1)) at higher temperature (43 °C) by this mutant could decrease the overall cost of fermentation process and increase the quality of ethanol production.

[Characteristics of extracellular invertase of Saccharomyces cerevisiae in Heterologous expression of the suc2 gene in Solarium Tuberosum plants].

Some properties and activity of extracellular invertase in the Saccharomyces cerevisiae yeasts encoded by the suc2 gene in heterologous expression were described. It was shown that the target suc2 gene is actively expressed in the genome of the transformed potato plants and S. cerevisiae invertase synthesized by this gene is transported into the apoplast due to the signal peptide of the proteinase II inhibitor. This enzyme is present in the apoplast in a soluble form and absorbed into the cell wall.