Model-based biotechnological potential analysis of Kluyveromyces marxianus central metabolism.

The non-conventional yeast Kluyveromyces marxianus is an emerging industrial producer for many biotechnological processes. Here, we show the application of a biomass-linked stoichiometric model of central metabolism that is experimentally validated, and mass and charge balanced for assessing the carbon conversion efficiency of wild type and modified K. marxianus. Pairs of substrates (lactose, glucose, inulin, xylose) and products (ethanol, acetate, lactate, glycerol, ethyl acetate, succinate, glutamate, phenylethanol and phenylalanine) are examined by various modelling and optimisation methods. Our model reveals the organism's potential for industrial application and metabolic engineering. Modelling results imply that the aeration regime can be used as a tool to optimise product yield and flux distribution in K. marxianus. Also rebalancing NADH and NADPH utilisation can be used to improve the efficiency of substrate conversion. Xylose is identified as a biotechnologically promising substrate for K. marxianus.

Production of extracellular fructans by Gluconobacter nephelii P1464.

UNLABELLED: Bacterial extracellular fructans, known as levans, have potential applications in food, pharmaceutical and cosmetic industries and high fructan producing strains could contribute into the cost reduction and more extensive commercial usage of them. An acetic acid bacteria (AAB) isolate P1464 was obtained from the Microbial Strain Collection of Institute of Microbiology and Biotechnology, University of Latvia and identified as Gluconobacter nephelii by DNA-DNA hybridization and the formation of extracellular fructans by this strain was confirmed. Isolated extracellular fructose polymers were characterized using FT-IR spectroscopy and the structural features of fructan appeared as similar to the reference sample of bacterial levan. Molecular mass estimates showed that the isolated G. nephelii P1464 fructose polymer has a relatively small molecular weight (Mw 1122·939 ± 153·453 kDa) and a sizeable polydispersity (Mw/Mn = 21·57 ± 1·60), as compared with other AAB, which could promote their physiological activity, including the prebiotic effects. Obtained at different cultivation conditions characteristics of fructan production, including the biotechnological indices such as the productivity (Qp) and yield (Yp/s) ranging from 0·774 to 1·244 g l(-1)  h and from 0·181 to 0·436 g g(-1) , respectively, confirmed, that G. nephelii P1464 could be used as promising strain for commercial production of levan. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacterial fructans, known as levans, have extensive options for practical usage, however, actually limited due to high production costs. Therefore, the searches for efficient producer strains should be an urgent task to reduce costs. This study is the first report on the formation of fructans by a novel strain of acetic acid bacteria (AAB) Gluconobacter nephelii P1464. Characteristics obtained at different cultivation conditions confirmed the operation of a competitive and perspective producer strain. Isolated extracellular fructans are characterized by a lower molecular weight as compared with other AAB which could promote their physiological activity, including the prebiotic effects.

The effect of osmo-induced stress on product formation by Zymomonas mobilis on sucrose.

The intensification of biosynthesis of fructooligosaccharides in the presence of high salt concentrations was observed during sucrose (10%) fermentation by Zymomonas mobilis 113S. A 0.6 M NaCl concentration led to an increase of oligosaccharide productivity by 3.5-fold. Sorbitol formation was increased in the presence of 0.16 M NaCl and was inhibited at highest salt concentrations. In a medium with high (65%, w/w) sucrose content the salts gave inhibitory effects on fructooligosaccharide production by lyophilised Z. mobilis cells. Influence of salts on gluconic acid and sorbitol formation under these conditions was studied. The ratio of oligosaccharides and gluconic acid productivity (Qolig./Qglucon.) was increased approximately 2 times at 1% KCl. Sorbitol formation was not significantly influenced in the presence of KCl (up to 2%).

An elevation of the molar growth yield of Zymomonas mobilis during aerobic exponential growth.

Elevated values of molar growth yield (Yx/s = 14-26 g mol-1) were obtained during exponential growth (mu > 0.4 h-1) of Zymomonas mobilis ATCC 29191 by using reduced concentrations of glucose (6. 25-100 mM) and increased oxygen supply (Eh > 300 mV) in the growth medium, as compared to the Yx/s of anaerobic exponential growth (8-10 g mol-1). Aerobically grown cells showed an increased maximum growth rate (mumax), and a reduced specific glucose consumption rate (qs), and specific ethanol formation rate (qp), thus demonstrating a more pronounced energy-coupling growth under oxic conditions. These results can be neither explained by the concept of a solely operating Entner-Doudoroff pathway as an ATP source in aerobically growing cultures of Z. mobilis nor considered to be consistent with existing data on the lack of the Pasteur effect in this bacterium. Therefore, the results rather give evidence for the essential contribution of aerobic ATP generation under the reported conditions.

Indolylacetic acid and N6-(delta 2-isopentenyl) adenine affect NADH binding to yeast alcohol dehydrogenase and inhibit in vitro the enzymatic oxidation of ethanol.

Derivative UV spectroscopic data show that the plant growth substances N6-(delta 2-isopentenyl) adenine (i6Ade) and indolylacetic acid (IAA) can bind to the yeast alcohol dehydrogenase (ADH) and affect coenzyme-enzyme binding. This is confirmed by enzyme kinetics studies. At fixed ethanol concentrations (27.8 and 111.1 mM) and varying NAD+ concentrations (0.033-2 mM), as well as at fixed levels of coenzyme (0.67 and 2 mM), and at varying concentrations of ethanol (1.4-111.1 mM), the rate of ethanol oxidation is significantly inhibited by i6Ade and IAA. The kinetics of the ADH reaction is affected by two inhibition constants (KI and K'I) which correspond to the dissociation constants of complexes EI and ESI, respectively. For i6Ade the KI = 0.52 +/- 0.06 mM and K'I = 0.74 +/- 0.07 mM, and for IAA the KI = 0.88 +/- 0.03 mM and K'I = 0.99 +/- 0.02 mM.

[Stability to the extreme influences and significance of internal pH of Saccharomyces cerevisiae depending on the density of cellular suspensions]. / Ustoichivost' k ékstremal'nym vozdeistviiam i znachenie vnutrennego pH Saccharomyces cerevisiae v zavisimost ot plotnosti kletochnykh suspenzii.

The resistance of a Saccharomyces cerevisiae population against gamma-irradiation (400 krd), heating (60 degrees C), repeated freezing--thawing and rehydration of dehydrated cells was shown to depend on the density of a cell suspension. The percentage of damaged cells (stained with primulin) was found to increase in a suspension with decreasing cell concentrations. The effect was also exerted in an oxygen-free medium. The percentage of undamaged cells increased in populations with a lower cell concentration when a more viscous medium (0.2% agar) was used. The intracellular pH dropped down in a denser intact yeast suspension and rose with an increase in the concentration of dehydrated-rehydrated cells.

[Distribution of trehalose between the cells and the rehydration medium in dehydrated Saccharomyces cerevisiae]. / Raspredelenie tregalozy mezhdu kletkami i sredoi regidratatsii u obezvozhennykh Saccharomyces cerevisiae.

The work was concerned with studying the balance of trehalose distribution between the rehydration medium and Saccharomyces cerevisiae cells grown in a chemically defined medium and dehydrated using the convective technique. A direct linear correlation between the viability of populations and the overall residual trehalose content in the cells and in the medium after the rehydration of dry yeast cells was shown to be most important. An inverse correlation was established between the viability of yeast cells and the amount of trehalose mobilised by the cells in the process of rehydration.

[Rehydration conditions for dehydrated Saccharomyces cerevisiae yeasts and cell viability]. / Usloviia regidratatsii obezvozhennykh drozhzhei Saccharomyces cerevisiae i zhiznesposobnost' kletok.

The viability of dehydrated Saccharomyces cerevisiae populations was studied as a result of changes in the conditions of rehydration (temperature, pH, time, composition of the medium). Rehydration was shown to play a key role for the functional characteristics of populations in the general consideration of dehydration-rehydration and to involve several factors. The viability of cells is regarded as both a relative and absolute criterion for the physiological state of populations. The conditions of rehydration should be strictly standardized for dehydrated microorganisms in comprehensive studies of mechanisms involved in the stability of microorganisms.

[Dehydration effect on the fatty acid composition of lipids in baker's yeast Saccharomyces cerevisiae 14]. / Vliianie obezvozhivaniia na zhirnokislotnyi sostav lipidov drozhzhei Saccharomyces cerevisiae.

The effect of dehydration on the fatty acid composition of lipids in baker's yeast Saccharomyces cerevisiae 14 cultivated on the ethanol (1.5%) containing medium in the presence or absence of glutamic acid and biotin or on molasses (2%) was studied. In spite of a relatively low drying temperature (35 degrees C), the correlation between saturated and non-saturated fatty acids may vary within 40--60 min.