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1.
Petrus Jakobus van Zyl Stephanus Gouws Kilian Bernard Alexander Prior 《Applied microbiology and biotechnology》1993,39(2):235-241
When cell suspensions of Zygosaccharomyces rouxii were subjected to osmotic shock with NaCl, the cell volume decreased sharply and plasmolysis was observed. The cell subsequently recovered and volumes similar to those of cells growing at the respective water activity (aw) values were found. Cycloheximide prevented cell recovery, indicating the involvement of protein synthesis in the recovery process. The intracellular glycerol concentration of Z. rouxii incubated in the presence of [14C]glycerol increased from 13 to 96 mmol/l during the initial 20 min after an upshock from 0.998 aw to 0.96 aw. All the intracellular glycerol was labelled and therefore derived from the medium. Labelled glycerol was subsequently utilized and replaced by unlabelled glycerol produced by the cell within 90 min. The initial increase in glycerol concentration following the upshock was confirmed by 13C-nuclear magnetic resonance (NMR) spectroscopic studies of cell extracts. The combined dihydroxyacetone and dihydroxyacetone phosphate concentrations fluctuated during this period, whereas glycerol-3-phosphate initially increased and then remained constant. This indicates that the production of glycerol is regulated. Decreases in ATP and polyphosphate levels were observed following osmotic upshock and may reflect a greater demand for ATP during the period of adjustment to decreased aw. The changes in cell volume and in ATP concentration following osmotic upshock may serve as osmoregulatory signals in Z. rouxii, as suggested previously for other microorganisms.
Correspondence to: S. G. Kilian 相似文献
2.
Petrus Jakobus van Zyl Bernard Alexander Prior 《Applied microbiology and biotechnology》1990,33(1):12-17
Summary Glycerol and arabitol were the main polyols accumulated by Zygosaccharomyces rouxii in continuous culture but the intracellular and extracellular concentrations of the polyols varied with the dilution rate and osmoticum used to adjust the water activity (aw) to 0.960. When the aw was adjusted with NaCl, glycerol was the main polyol accumulated intracellularly whereas glycerol and arabitol were accumulated when polyethylene glycol (PEG) 400 was used. The extracellular glycerol and arabitol concentrations at 0.960 aw (NaCl or PEG 400) were similar or decreased relative to cultures at 0.998 aw. Compared to steady-state cultivation at 0.998 aw, the yeast retained at 0.960 aw (NaCl or PEG 400) a greater proportion of the total glycerol intracellularly against an increased concentration ratio without significantly greater production of glycerol. Arabitol was only significant in osmoregulation when cultivated at 0.960 aw (PEG 400). The intracellular glycerol concentration was insufficient to balance the aw across the membrane, but an equilibrium could be achieved under certain conditions if arabitol was also osmotically active.
Offprint requests to: P. J. van Zyl 相似文献
3.
Glycerol formation is vital for reoxidation of nicotinamide adenine dinucleotide (reduced form; NADH) under anaerobic conditions
and for the hyperosmotic stress response in the yeast Saccharomyces cerevisiae. However, relatively few studies have been made on hyperosmotic stress under anaerobic conditions. To study the combined
effect of salt stress and anaerobic conditions, industrial and laboratory strains of S. cerevisiae were grown anaerobically on glucose in batch-cultures containing 40 g/l NaCl. The time needed for complete glucose conversion
increased considerably, and the specific growth rates decreased by 80–90% when the cells were subjected to the hyperosmotic
conditions. This was accompanied by an increased yield of glycerol and other by-products and reduced biomass yield in all
strains. The slowest fermenting strain doubled its glycerol yield (from 0.072 to 0.148 g/g glucose) and a nearly fivefold
increase in acetate formation was seen. In more tolerant strains, a lower increase was seen in the glycerol and in the acetate,
succinate and pyruvate yields. Additionally, the NADH-producing pathway from acetaldehyde to acetate was analysed by overexpressing
the stress-induced gene ALD3. However, this had no or very marginal effect on the acetate and glycerol yields. In the control experiments, the production
of NADH from known sources well matched the glycerol formation. This was not the case for the salt stress experiments in which
the production of NADH from known sources was insufficient to explain the formed glycerol. 相似文献
4.
Saburova E. A. Avseenko N. V. Simonova N. B. Elfimova L. I. Pronina N. A. Semenenko V. E. 《Russian Journal of Plant Physiology》2000,47(6):761-771
The dependence of the catalytic properties of lactate dehydrogenase (LDH, EC 1.1.1.27) from a halophilic alga Dunaliella salina, a glycophilic alga Chlamydomonas reinhardtii, and from porcine muscle on glycerol concentration, medium pH, and temperature was investigated. Several chemical properties of the enzyme from D. salina differentiated it from the LDH preparation obtained from C. reinhardtii and any homologous enzymes of plant, animal, and bacterial origin. (1) V
max of pyruvate reduction manifested low sensitivity to the major intracellular osmolyte, glycerol. (2) The affinity of LDH for its coenzyme NADH dropped in the physiological pH region of 6–8. Above pH 8, NADH virtually did not bind to LDH, while the enzyme affinity for pyruvate did not change considerably. (3) The enzyme thermostability was extremely low: LDH was completely inactivated at room temperature within 30 min. The optimum temperature for pyruvate reduction (32°C) was considerably lower than with the enzyme preparations from C. reinhardtii (52°C) and porcine muscle (61°C). (4) NADH greatly stabilized LDH: the ratio of LDH inactivation constants in the absence of the coenzyme and after NADH addition at the optimum temperature in the preparation from D. salina exceeded the corresponding indices of LDH preparations from C. reinhardtii twelve times and from porcine muscle eight times. The authors believe that these LDH properties match the specific metabolism of D. salina which is set at rapid glycerol synthesis under hyperosmotic stress conditions. The increase of cytoplasmic pH value produced in D. salina by the hyperosmotic shock can switch off the terminal reaction of the glycolytic pathway and thus provide for the most efficient utilization of NADH in the cycle of glycerol synthesis. As LDH is destabilized in the absence of NADH, this reaction is also switched off. In the course of alga adaptation to the hyperosmotic shock, glycerol accumulation and the neutralization of intracellular pH stabilize LDH, thus creating the conditions for restoring the complete glycolytic cycle. 相似文献
5.
Lei Fang Hui Xin Lin Chin Seng Low Mei Hui Wu Yvonne Chow Yuan Kun Lee 《Plant biotechnology journal》2012,10(9):1129-1135
Bioengineering of photoautotrophic microalgae into CO2 scrubbers and producers of value‐added metabolites is an appealing approach in low‐carbon economy. A strategy for microalgal bioengineering is to enhance the photosynthetic carbon assimilation through genetically modifying the photosynthetic pathways. The halotolerant microalgae Dunaliella posses an unique osmoregulatory mechanism, which accumulates intracellular glycerol in response to extracellular hyperosmotic stresses. In our study, the Calvin cycle enzyme sedoheptulose 1,7‐bisphosphatase from Chlamydomonas reinhardtii (CrSBPase) was transformed into Dunaliella bardawil, and the transformant CrSBP showed improved photosynthetic performance along with increased total organic carbon content and the osmoticum glycerol production. The results demonstrate that the potential of photosynthetic microalgae as CO2 removers could be enhanced through modifying the photosynthetic carbon reduction cycle, with glycerol as the carbon sink. 相似文献
6.
Tennant-Eyles AJ Moffitt H Whitehouse CA Roberts RG 《Biochemical and biophysical research communications》2011,410(3):471-477
Myo-inositol is one of the major organic osmolytes in the brain and the kidney. The accumulation of intracellular organic osmolytes allows cells to regulate intracellular osmolality without altering cytoplasmic ionic strength and to adapt to hyperosmotic conditions. Two types of myo-inositol transporters, sodium/myo-inositol transporter and H+/myo-inositol transporter (HMIT), have been identified. Sodium/myo-inositol transporters are induced by osmotic stress and might be involved in the intracellular accumulation of myo-inositol in mammals. The role of HMIT, however, remains unknown. In the present study, we characterized three Caenorhabditis elegansHMIT genes, hmit-1.1, hmit-1.2, and hmit-1.3. hmit-1.1 was expressed in the intestine, and hmit-1.2 was expressed in the glia and the excretory canal, which is an osmotic regulatory organ that is functionally analogous to the kidney. hmit-1.3 was expressed in the intestine and the glia. The expression of hmit-1.1 and hmit-1.2 but not hmit-1.3, was markedly induced under hyperosmotic conditions. Animals with mutant hmit-1.1 and hmit-1.2 were hypersensitive to osmotic stress. The defects of hmit-1.1 and hmit-1.2 mutants were rescued by hmit-1.1 and hmit-1.2 transgenes, respectively, and by modified human HMIT. In human cell lines, HMIT expression was induced in hyperosmotic conditions. These findings indicate that the C. elegans HMIT family has a crucial role in the osmoprotective response. 相似文献
7.
A soluble NAD-dependent alcohol dehydrogenase (ADH) activity was detected in mycelium and yeast cells of wild-type Mucor rouxii. In the mycelium of cells grown in the absence of oxygen, the enzyme activity was high, whereas in yeast cells, ADH activity
was high regardless of the presence or absence of oxygen. The enzyme from aerobically or anaerobically grown mycelium or yeast
cells exhibited a similar optimum pH for the oxidation of ethanol to acetaldehyde (∼pH 8.5) and for the reduction of acetaldehyde
to ethanol (∼pH 7.5). Zymogram analysis conducted with cell-free extracts of the wild-type and an alcohol-dehydrogenase-deficient
mutant strain indicated the existence of a single ADH enzyme that was independent of the developmental stage of dimorphism,
the growth atmosphere, or the carbon source in the growth medium. Purified ADH from aerobically grown mycelium was found to
be a tetramer consisting of subunits of 43 kDa. The enzyme oxidized primary and secondary alcohols, although much higher activity
was displayed with primary alcohols. K
m values obtained for acetaldehyde, ethanol, NADH2, and NAD+ indicated that physiologically the enzyme works mainly in the reduction of acetaldehyde to ethanol.
Received: 11 March 1999 / Accepted: 14 July 1999 相似文献
8.
9.
In response to osmotic stress, proline is accumulated in many bacterial and plant cells. During various stresses, the yeast
Saccharomyces cerevisiae induces glycerol or trehalose synthesis, but the fluctuations in gene expression and intracellular levels of proline in yeast
are not yet well understood. We previously found that proline protects yeast cells from damage by freezing, oxidative, or
ethanol stress. In this study, we examined the relationships between the gene expression profiles and intracellular contents
of glycerol, trehalose, and proline under stress conditions. When yeast cells were exposed to 1 M sorbitol stress, the expression
of GPD1 encoding glycerol-3-phosphate dehydrogenase is induced, leading to glycerol accumulation. In contrast, in the presence of
9% ethanol, the rapid induction of TPS2 encoding trehalose-6-phosphate phosphatase resulted in trehalose accumulation. We found that intracellular proline levels
did not increase immediately after addition of sorbitol or ethanol. However, the expressions of genes involved in proline
synthesis and degradation did not change during exposure to these stresses. It appears that the elevated proline levels are
due primarily to an increase in proline uptake from a nutrient medium caused by the induction of PUT4. These results suggest that S. cerevisiae cells do not accumulate proline in response to sorbitol or ethanol stress different from other organisms. 相似文献
10.
Minoska Valli Michael Sauer Paola Branduardi Nicole Borth Danilo Porro Diethard Mattanovich 《Applied microbiology》2005,71(3):1515-1521
Intracellular pH has an important role in the maintenance of the normal functions of yeast cells. The ability of the cell to maintain this pH homeostasis also in response to environmental changes has gained more and more interest in both basic and applied research. In this study we describe a protocol which allows the rapid determination of the intracellular pH of Saccharomyces cerevisiae cells. The method is based on flow cytometry and employs the pH-dependent fluorescent probe carboxy SNARF-4F. The protocol attempts to minimize the perturbation of the system under study, thus leading to accurate information about the physiological state of the single cell. Moreover, statistical analysis performed on major factors that may influence the final determination supported the validity of the optimized protocol. The protocol was used to investigate the effect of external pH on S. cerevisiae cells incubated in buffer. The results obtained showed that stationary cells are better able than exponentially grown cells to maintain their intracellular pH homeostasis independently of external pH changes. Furthermore, analysis of the intracellular pH distribution within the cell populations highlighted the presence of subpopulations characterized by different intracellular pH values. Notably, a different behavior was observed for exponentially grown and stationary cells in terms of the appearance and development of these subpopulations as a response to a changing external pH. 相似文献
11.
Aerobic mycelium of wild-type Mucor rouxii accumulated about ten times higher amounts of the carotenoid pigment -carotene when grown continuously in the presence of light than the corresponding cultures grown in the dark. Carotenoid accumulation was dependent on light intensity, with the threshold located at about 10-2 W.m-2. Photocarotenogenesis in complex medium was more efficient with glucose as a carbon source. Carotenoid synthesis by M. rouxii mycelium was unaffected by both retinol acetate and retinal, which are stimulators of carotenogenesis in other zygomycetes. Carotenogenesis was significant in aerobic mycelium but was almost undetectable in anaerobic mycelium as well as in aerobic or anaerobic yeast cells. This suggested an involvement of oxygen in carotenoid synthesis by M. rouxii and the existence of developmental regulation of the expression or operation of the pathway. 相似文献
12.
Myo-inositol (Ins) is a major compatible osmolyte in many cells, including those of Mozambique tilapia (Oreochromis mossambicus). Ins biosynthesis is highly up-regulated in tilapia and other euryhaline fish exposed to hyperosmotic stress. In this study, enzymatic regulation of two enzymes of Ins biosynthesis, Ins phosphate synthase (MIPS) and inositol monophosphatase (IMPase), by direct ionic effects is analyzed. Specific MIPS and IMPase isoforms from Mozambique tilapia (MIPS-160 and IMPase 1) were selected based on experimental, phylogenetic, and structural evidence supporting their role for Ins biosynthesis during hyperosmotic stress. Recombinant tilapia IMPase 1 and MIPS-160 activity was assayed in vitro at ionic conditions that mimic changes in the intracellular milieu during hyperosmotic stress. The in vitro activities of MIPS-160 and IMPase 1 are highest at alkaline pH of 8.8. IMPase 1 catalytic efficiency is strongly increased during hyperosmolality (particularly for the substrate D-Ins-3-phosphate, Ins-3P), mainly as a result of [Na+] elevation. Furthermore, the substrate-specificity of IMPase 1 towards D-Ins-1-phosphate (Ins-1P) is lower than towards Ins-3P. Because MIPS catalysis results in Ins-3P this results represents additional evidence for IMPase 1 being the isoform that mediates Ins biosynthesis in tilapia. Our data collectively demonstrate that the Ins biosynthesis enzymes are activated under ionic conditions that cells are exposed to during hypertonicity, resulting in Ins accumulation, which, in turn, results in restoration of intracellular ion homeostasis. We propose that the unique and direct ionic regulation of the activities of Ins biosynthesis enzymes represents an efficient biochemical feedback loop for regulation of intracellular physiological ion homeostasis during hyperosmotic stress. 相似文献
13.
Saha BC Sakakibara Y Cotta MA 《Journal of industrial microbiology & biotechnology》2007,34(7):519-523
A newly isolated Zygosaccharomyces rouxii NRRL 27,624 produced d-arabitol as the main metabolic product from glucose. In addition, it also produced ethanol and glycerol. The optimal conditions
were temperature 30°C, pH 5.0, 350 rpm, and 5% inoculum. The yeast produced 83.4 ± 1.1 g d-arabitol from 175 ± 1.1 g glucose per liter at pH 5.0, 30°C, and 350 rpm in 240 h with a yield of 0.48 g/g glucose. It also
produced d-arabitol from fructose, galactose, and mannose. The yeast produced d-arabitol and xylitol from xylose and also from a mixture of xylose and xylulose. Resting yeast cells produced 63.6 ± 1.9 g
d-arabitol from 175 ± 1.8 g glucose per liter in 210 h at pH 5.0, 30°C and 350 rpm with a yield of 0.36 g/g glucose. The yeast
has potential to be used for production of xylitol from glucose via d-arabitol route.
Mention of trade names or commercial products in this article is solely for the purpose of providing specific information
and does not imply recommendation or endorsement by the U.S. department of Agriculture. 相似文献
14.
Petrus Jakobus van Zyl Bernard Alexander Prior Stephanus Gouws Kilian 《Applied microbiology and biotechnology》1991,36(3):369-374
Summary Enzyme analyses indicated that the metabolism of glycerol by Zygosaccharomyces rouxii occurred via either glycerol-3-phosphate (G3P) or dihydroxyacetone (DHA). The route via DHA is significant in osmoregulation. The specific activities of glycerol dehydrogenase (GDHG) and DHA kinase, which metabolize glycerol via DHA, increased nine- and fourfold respectively during osmotic stress [0.960 water activity (aw) adjusted with NaCl] when compared to non-stressed conditions (0.998 aw). Both pathways are under metabolic regulation. Glycerol kinase, mitochondrial G3P dehydrogenase and DHA kinase are induced by glycerol while the latter is also repressed by glucose. Cells treated with cycloheximide prior to osmotic upshock showed significantly lower DHA kinase and GDHG levels and lower intracellular glycerol concentrations when compared to untreated control cells. Thus protein synthesis is essential for osmotic adaptation.
Offprint requests to: B. A. Prior 相似文献
15.
Malonyl-CoA is an essential precursor for fatty acid biosynthesis that is generated from the carboxylation of acetyl-CoA.
In this work, a gene coding for acetyl-CoA carboxylase (ACC) was isolated from an oleaginous fungus, Mucor rouxii. According to the amino acid sequence homology and the conserved structural organization of the biotin carboxylase, biotin
carboxyl carrier protein, and carboxyl transferase domains, the cloned gene was characterized as a multi-domain ACC1 protein.
Interestingly, a 40% increase in the total fatty acid content of the non-oleaginous yeast Hansenula polymorpha was achieved by overexpressing the M. rouxii ACC1. This result demonstrated a significant improvement in the production of fatty acids through genetic modification in
this yeast strain. 相似文献
16.
The yeast Candida bombicola (ATCC 22214) grown on primary carbon source glucose (100 g l−1) and secondary carbon, arachidonic acid (2 g l−1) produced mixture of sophorolipids up to 1.44 g l−1. The crude product was a heterogeneous mixture of sophorolipids, which are glycolipids of sophorose linked to the fatty acid
through glycosidic bond between ω and ω−1 carbon of arachidonic acid. The derived sophorolipids were isolated by silica gel
chromatography using dialysis tubing. The purified sophorolipids were characterized by ESI-MS and FT-IR. Acid hydrolysis of
the resolved sophorolipids were characterized by ESI-MS for the presence of 20-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid (20-HETE) and 19-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid (19-HETE), compounds of pronounced pharmaceutical importance. 相似文献
17.
Yeast identification in grape juice concentrates from Argentina 总被引:1,自引:0,他引:1
Combina M Daguerre C Massera A Mercado L Sturm ME Ganga A Martinez C 《Letters in applied microbiology》2008,46(2):192-197
Aims: The purpose of this study was to identify yeast species present in spoiled and unspoiled grape juice concentrates from Argentine industries. Methods and Results: Osmophilic and osmotolerant yeasts were isolated from spoiled – visually effervescent – and unspoiled – without any visible damage – grape juice concentrates by the spread‐plate technique in two culture media. Yeast identification was done by classical and molecular methods. Zygosaccharomyces rouxii was the only species isolated from spoiled grape juice concentrates. In unspoiled samples, five different species were identified: Z. rouxii was isolated at a higher frequency, followed in decreasing order by Saccharomyces cerevisiae, Schizosaccharomyces pombe, Pichia anomala and Kluyveromyces delphensis. Conclusions: Yeasts isolated from grape juice concentrates were characterized by a limited taxonomic diversity, where Z. rouxii was the main species isolated. Significance and Impact of the Study: Grape production in Argentina is mainly devoted to the industry where wine and grape juice concentrates represent major types of commercial products. Little information on common yeast contaminants is available for grape juice concentrates. This study constitutes the first report of osmophilic yeast species present in spoiled and unspoiled grape juice concentrates elaborated in Argentina. 相似文献
18.
Tomohiko Matsuzawa Takao Ohashi Akira Hosomi Naotaka Tanaka Hideki Tohda Kaoru Takegawa 《Applied microbiology and biotechnology》2010,87(2):715-727
The budding yeast Saccharomyces cerevisiae is able to utilize glycerol as the sole carbon source via two pathways (glycerol 3-phosphate pathway and dihydroxyacetone
[DHA] pathway). In contrast, the fission yeast Schizosaccharomyces pombe does not grow on media containing glycerol as the sole carbon source. However, in the presence of other carbon sources such
as galactose and ethanol, S. pombe could assimilate glycerol and glycerol was preferentially utilized over ethanol and galactose. No equivalent of S. cerevisiae Gcy1/glycerol dehydrogenase has been identified in S. pombe. However, we identified a gene in S. pombe, SPAC13F5.03c (gld1
+), that is homologous to bacterial glycerol dehydrogenase. Deletion of gld1 caused a reduction in glycerol dehydrogenase activity and prevented glycerol assimilation. The gld1Δ cells grew on 50 mM DHA as the sole carbon source, indicating that the glycerol dehydrogenase encoded by gld1
+ is essential for glycerol assimilation in S. pombe. Strains of S. pombe deleted for dak1
+ and dak2
+ encoding DHA kinases could not grow on glycerol and showed sensitivity to a higher concentration of DHA. The dak1Δ strain showed a more severe reduction of growth on glycerol and DHA than the dak2Δ strain because the expression of dak1
+ mRNA was higher than that of dak2
+. In wild-type S. pombe, expression of the gld1
+, dak1
+, and dak2
+ genes was repressed at a high concentration of glucose and was derepressed during glucose starvation. We found that gld1
+ was regulated by glucose repression and that it was derepressed in scr1Δ and tup12Δ strains. 相似文献
19.
Functional interactions between potassium and phosphate homeostasis in Saccharomyces cerevisiae
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David Canadell Asier González Carlos Casado Joaquín Ariño 《Molecular microbiology》2015,95(3):555-572
Maintenance of ion homeostatic mechanisms is essential for living cells, including the budding yeast Saccharomyces cerevisiae. Whereas the impact of changes in phosphate metabolism on metal ion homeostasis has been recently examined, the inverse effect is still largely unexplored. We show here that depletion of potassium from the medium or alteration of diverse regulatory pathways controlling potassium uptake, such as the Trk potassium transporters or the Pma1 H+‐ATPase, triggers a response that mimics that of phosphate (Pi) deprivation, exemplified by accumulation of the high‐affinity Pi transporter Pho84. This response is mediated by and requires the integrity of the PHO signaling pathway. Removal of potassium from the medium does not alter the amount of total or free intracellular Pi, but is accompanied by decreased ATP and ADP levels and rapid depletion of cellular polyphosphates. Therefore, our data do not support the notion of Pi being the major signaling molecule triggering phosphate‐starvation responses. We also observe that cells with compromised potassium uptake cannot grow under limiting Pi conditions. The link between potassium and phosphate homeostasis reported here could explain the invasive phenotype, characteristic of nutrient deprivation, observed in potassium‐deficient yeast cells. 相似文献
20.
In this study Banh mensamples obtained from Vietnam were analysed in terms of pH, moisture content and fungal composition. The banh men pH proved to be acidic with a mean pH of 5.76. Moisture content was 13.6%. Total mould and yeast counts yielded 1.3 × 106 and 4.3 × 106 c.f.u./g-fresh sample, respectively. A total of 53 fungal isolates were obtained from 20 moulds and 33 yeasts. The mould isolates were identified as Rhizopus oryzae, Mucor indicus, Mucor circinilloides, and Amylomyces rouxii. The yeast isolates were identified as Saccharomyces cerevisiae, Hyphopichia burtonii, Saccharomycopsis fibuligera, Pichia anomala, and Candida sp. Based on the parameters used in this study, it can be deduced that banh men is similar to ragi and other Asian fermentation starters. 相似文献