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Trehalose
Kinds of Trehalose Terms modified by Trehalose Selected AbstractsGUSTATORY REACTION TIME AND TIME INTENSITY MEASUREMENTS OF TREHALOSE AND SUCROSE SOLUTIONS AND THEIR MIXTURESJOURNAL OF SENSORY STUDIES, Issue 2 2009MARA VIRGINIA GALMARINI ABSTRACT Dynamic sweetness perception of commercial food grade trehalose, sucrose solutions and their mixtures were studied for a wide range of concentrations. For gustatory reaction time (GRT), concentrations ranged from 2.3 to 13.8% for sucrose and up to 23.0% for trehalose. For time intensity (T-I) sucrose or trehalose solutions (concentration range 2.3,36.8%) and their combinations (23.0 and 36.8% total solids) were analyzed. Trehalose had bigger GRT along the studied range. Both sugars presented similar values for persistence and times of plateau and to maximum intensity, while a significant difference was observed in intensity and GRT at equal concentrations. Trehalose had longer persistence than sucrose in equi -sweet solutions. Overall sweetness profile of some sucrose solutions (i.e., 29.9% sucrose solution and 0.6 sucrose/trehalose ratio mixture at 36.8% total solids) were perceived as similar to mixtures of sucrose/trehalose or single trehalose solutions, which suggests the possibility of sugar replacement without completely modifying sweetness perception. PRACTICAL APPLICATIONS It has been suggested that trehalose may be a potential substitute for sucrose and other sugars used in food formulation because, although its chemical structure is very similar to that of sucrose, it is more stable at low pH and high temperatures. It is not involved in caramelization and does not participate in Maillard reaction with amino acids/proteins. In order to fully establish the potential of trehalose as a functional replacement of sucrose we have determined the sweetness dynamic profile (gustatory reaction time and time-intensity curves) of trehalose solutions and sucrose/trehalose solutions; this aspect is needed for adequately replacing (partially or totally) sucrose in food systems. [source] Development of Ketoside-Type Analogues of Trehalose by Using ,-Stereoselective O-Glycosidation of KetoseEUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 22 2007Rie Namme Abstract The stereoselective synthesis of ketoside-type analogues of trehalose is described. O-Glycosidation of hept-2-ulopyranose with trimethylsilyl ,-pyranoside promoted by trimethylsilyl trifluoromethanesulfonate afforded ,-ketopyranosyl ,-aldopyranosides exclusively. ,-Ketopyranosyl ,-aldooyranosides and ,-ketopyranosyl ,-ketopyranosides were also synthesized in a similar manner. The benzyl protecting groups of the hydroxy moieties were removed by hydrogenolysis to afford fully deprotected trehalose analogues.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Enzymatic control of anhydrobiosis-related accumulation of trehalose in the sleeping chironomid, Polypedilum vanderplankiFEBS JOURNAL, Issue 20 2010Kanako Mitsumasu Larvae of an anhydrobiotic insect, Polypedilum vanderplanki, accumulate very large amounts of trehalose as a compatible solute on desiccation, but the molecular mechanisms underlying this accumulation are unclear. We therefore isolated the genes coding for trehalose metabolism enzymes, i.e. trehalose-6-phosphate synthase (TPS) and trehalose-6-phosphate phosphatase (TPP) for the synthesis step, and trehalase (TREH) for the degradation step. Although computational prediction indicated that the alternative splicing variants (PvTps,/,) obtained encoded probable functional motifs consisting of a typical consensus domain of TPS and a conserved sequence of TPP, PvTps, did not exert activity as TPP, but only as TPS. Instead, a distinct gene (PvTpp) obtained expressed TPP activity. Previous reports have suggested that insect TPS is, exceptionally, a bifunctional enzyme governing both TPS and TPP. In this article, we propose that TPS and TPP activities in insects can be attributed to discrete genes. The translated product of the TREH ortholog (PvTreh) certainly degraded trehalose to glucose. Trehalose was synthesized abundantly, consistent with increased activities of TPS and TPP and suppressed TREH activity. These results show that trehalose accumulation observed during anhydrobiosis induction in desiccating larvae can be attributed to the activation of the trehalose synthetic pathway and to the depression of trehalose hydrolysis. [source] Trehalose and trehalose-hydrolyzing enzyme in the haemolymph of Locusta migratoria infected with Metarhizium anisopliae strain CQMa102INSECT SCIENCE, Issue 4 2007HUA ZHAO Abstract Topical application of the Metarhizium anisopliae var. acridum specialist strain CQMa102 to the locust Locusta migratoria manilensis results in changes of the concentrations of trehalose and glucose in the haemolymph. Micrographs of the locust haemolymph shows Metarhizium anisopliae can effectivly penetrate the external skeleton of locust and after 2 days infection, the hyphae body will appear in the haemolymph of infected insects. The time in decrease of trehalose concentration coincided with that in increase of trehalose-hydrolysing enzyme activity in the haemolymph of the fungus-infected insects. Overlay gel analysis indicated there was considerably more trehalose-hydrolysing activity in the haemolymph of locusts infected by fungus than in controls. A comparable isoform was identified in in vitro culture of the fungus, suggesting a fungal origin for the in vivo enzyme. Haemolymph trehalose decreased significantly during mycosis of locusts by M. anisopliae. All these results suggested that this fungus may take advantage of competing nutrient utilization against the insect by its trehalose-hydrolyzing enzyme secretion. It may provide fundamental knowledge for fungal pathogenesis. [source] State transitions and physicochemical aspects of cryoprotection and stabilization in freeze-drying of Lactobacillus rhamnosus GG (LGG)JOURNAL OF APPLIED MICROBIOLOGY, Issue 6 2008K.S. Pehkonen Abstract Aims:, The frozen and dehydrated state transitions of lactose and trehalose were determined and studied as factors affecting the stability of probiotic bacteria to understand physicochemical aspects of protection against freezing and dehydration of probiotic cultures. Methods and Results:,Lactobacillus rhamnosus GG was frozen (,22 or ,43°C), freeze-dried and stored under controlled water vapour pressure (0%, 11%, 23% and 33% relative vapour pressure) conditions. Lactose, trehalose and their mixture (1 : 1) were used as protective media. These systems were confirmed to exhibit relatively similar state transition and water plasticization behaviour in freeze-concentrated and dehydrated states as determined by differential scanning calorimetry. Ice formation and dehydrated materials were studied using cold-stage microscopy and scanning electron microscopy. Trehalose and lactose,trehalose gave the most effective protection of cell viability as observed from colony forming units after freezing, dehydration and storage. Enhanced cell viability was observed when the freezing temperature was ,43°C. Conclusions:, State transitions of protective media affect ice formation and cell viability in freeze-drying and storage. Formation of a maximally freeze-concentrated matrix with entrapped microbial cells is essential in freezing prior to freeze-drying. Freeze-drying must retain a solid amorphous state of protectant matrices. Freeze-dried matrices contain cells entrapped in the protective matrices in the freezing process. The retention of viability during storage seems to be controlled by water plasticization of the protectant matrix and possibly interactions of water with the dehydrated cells. Highest cell viability was obtained in glassy protective media. Significance and Impact of the Study:, This study shows that physicochemical properties of protective media affect the stability of dehydrated cultures. Trehalose and lactose may be used in combination, which is particularly important for the stabilization of probiotic bacteria in dairy systems. [source] Effect of trehalose on survival of Bradyrhizobium japonicum during desiccationJOURNAL OF APPLIED MICROBIOLOGY, Issue 3 2003J.G. Streeter Abstract Aims: A major reason for the ineffectiveness of legume inoculants in the field is the rapid death of rhizobia because of desiccation. The major purpose of this study was to identify conditions under which ,,, -trehalose would improve survival of Bradyrhizobium japonicum during desiccation. Methods and Results: Trehalose was added to cultures just prior to desiccation or was supplied to bacteria during the 6-day growth period. A wide variety of trehalose concentrations was tested. Trehalose added to cultures at the time of desiccation improved survival slightly, but trehalose loading during growth was much more effective in protection against desiccation. Growth of bacteria with 3 mmol l,1 trehalose increased trehalose concentration in cells by about threefold and increased survival of cells placed on soya bean [Glycine max (L.) Merr.] seeds by two- to four-fold after 2 or 24 h. Average of overall results indicate that growth of bacteria with trehalose in the medium resulted in a 294% increase in survival after 24 h of desiccation. The concentration of trehalose in cells was very highly correlated with survival of bacteria. When trehalose-loaded cells were suspended in buffer or water, 60,85% of cellular trehalose was lost in about 1 h and, in spite of these losses, survival during desiccation was not reduced. Conclusions: Accumulation of trehalose in the cytoplasm is critical to the survival of B. japonicum during desiccation. Increasing the periplasmic concentration of trehalose is also beneficial but is not so critical as the concentration of trehalose in the cytoplasm. Because B. japonicum cannot utilize trehalose as a carbon source, cells can be loaded with trehalose by providing the disaccharide during the growth period. Significance and Impact of the Study: Although it may not be practical to use trehalose as a carbon source in inoculant production, it may be possible to engineer greater trehalose accumulation in rhizobia. Trehalose concentration in cells should be a useful predictor of survival during desiccation. [source] Trehalose extends longevity in the nematode Caenorhabditis elegansAGING CELL, Issue 4 2010Yoko Honda Summary Trehalose is a disaccharide of glucose found in diverse organisms and is suggested to act as a stress protectant against heat, cold, desiccation, anoxia, and oxidation. Here, we demonstrate that treatment of Caenorhabditis elegans with trehalose starting from the young-adult stage extended the mean life span by over 30% without any side effects. Surprisingly, trehalose treatment starting even from the old-adult stage shortly thereafter retarded the age-associated decline in survivorship and extended the remaining life span by 60%. Demographic analyses of age-specific mortality rates revealed that trehalose extended the life span by lowering age-independent vulnerability. Moreover, trehalose increased the reproductive span and retarded the age-associated decrease in pharyngeal-pumping rate and the accumulation of lipofuscin autofluorescence. Trehalose also enhanced thermotolerance and reduced polyglutamine aggregation. These results suggest that trehalose suppressed aging by counteracting internal or external stresses that disrupt protein homeostasis. On the other hand, the life span-extending effect of trehalose was abolished in long-lived insulin/IGF-1-like receptor (daf-2) mutants. RNA interference-mediated inactivation of the trehalose-biosynthesis genes trehalose-6-phosphate synthase-1 (tps-1) and tps-2, which are known to be up-regulated in daf-2 mutants, decreased the daf-2 life span. These findings indicate that a reduction in insulin/IGF-1-like signaling extends life span, at least in part, through the aging-suppressor function of trehalose. Trehalose may be a lead compound for potential nutraceutical intervention of the aging process. [source] Crystallization Kinetics and X-ray Diffraction of Crystals Formed in Amorphous Lactose, Trehalose, and Lactose/Trehalose MixturesJOURNAL OF FOOD SCIENCE, Issue 5 2005Song Miao ABSTRACT: Effects of storage time and relative humidity on crystallization kinetics and crystal forms produced from freeze-dried amorphous lactose, trehalose, and a lactose/trehalose mixture were compared. Samples were exposed to 4 different relative water vapor pressure (RVP) (44.1%, 54.5%, 65.6%, 76.1%) environments at room temperature. Crystallization was observed from time-dependent loss of sorbed water and increasing intensities of peaks in X-ray diffraction patterns. The rate of crystallization increased with increasing storage humidity. Lactose crystallized as ,-lactose monohydrate, ,-anhydrous, and anhydrous forms of ,- and ,-lactose in molar ratios of 5:3 and 4:1 in lactose and lactose/trehalose systems. Trehalose seemed to crystallize as a mixture of trehalose dihydrate and anhydrate in trehalose and lactose/trehalose systems. The crystal forms in a mixture of lactose and trehalose did not seem to be affected by the component sugars, but crystallization of the component sugars was delayed. Time-dependent crystallization of lactose and trehalose in the lactose-trehalose mixture could be modeled using the Avrami equation. The results indicated that crystallization data are important in modeling of crystallization phenomena and predicting stability of lactose and trehalose-containing food and pharmaceutical materials. Keywords: crystallization, lactose, trehalose, crystal form, X-ray diffraction [source] GUSTATORY REACTION TIME AND TIME INTENSITY MEASUREMENTS OF TREHALOSE AND SUCROSE SOLUTIONS AND THEIR MIXTURESJOURNAL OF SENSORY STUDIES, Issue 2 2009MARA VIRGINIA GALMARINI ABSTRACT Dynamic sweetness perception of commercial food grade trehalose, sucrose solutions and their mixtures were studied for a wide range of concentrations. For gustatory reaction time (GRT), concentrations ranged from 2.3 to 13.8% for sucrose and up to 23.0% for trehalose. For time intensity (T-I) sucrose or trehalose solutions (concentration range 2.3,36.8%) and their combinations (23.0 and 36.8% total solids) were analyzed. Trehalose had bigger GRT along the studied range. Both sugars presented similar values for persistence and times of plateau and to maximum intensity, while a significant difference was observed in intensity and GRT at equal concentrations. Trehalose had longer persistence than sucrose in equi -sweet solutions. Overall sweetness profile of some sucrose solutions (i.e., 29.9% sucrose solution and 0.6 sucrose/trehalose ratio mixture at 36.8% total solids) were perceived as similar to mixtures of sucrose/trehalose or single trehalose solutions, which suggests the possibility of sugar replacement without completely modifying sweetness perception. PRACTICAL APPLICATIONS It has been suggested that trehalose may be a potential substitute for sucrose and other sugars used in food formulation because, although its chemical structure is very similar to that of sucrose, it is more stable at low pH and high temperatures. It is not involved in caramelization and does not participate in Maillard reaction with amino acids/proteins. In order to fully establish the potential of trehalose as a functional replacement of sucrose we have determined the sweetness dynamic profile (gustatory reaction time and time-intensity curves) of trehalose solutions and sucrose/trehalose solutions; this aspect is needed for adequately replacing (partially or totally) sucrose in food systems. [source] Formulation of aqueous concentrated alumina suspensions.POLYMER INTERNATIONAL, Issue 4 2003Influence of a disaccharide: trehalose Abstract Trehalose, a disaccharide, is very well known to protect living cells efficiently from dehydration and has been tested in the formulation of aqueous alumina suspensions. It has been added into slurries stabilized with Tiron®, (HO)2C6H2(SO3Na)2, which permits a high state of dispersion by creating repulsive potential between particles. Trehalose added into such suspensions acts as a lubricant and enhances flowing properties of suspensions because of its strong interaction with water which breaks the hydrogen-bonded network of the solvent. Trehalose addition is beneficial for carrying out shaping methods of alumina components by coagulation as it increases solid concentration in the suspension, which facilitates pouring the suspension into a mould. Unfortunately this addition hinders coagulation of particles. © 2003 Society of Chemical Industry [source] Spectroscopic study of the physical properties making trehalose a stabilizing and shelf life extending compound in food industryQUALITY ASSURANCE & SAFETY OF CROPS & FOOD, Issue 2 2010S. Magazù Abstract Introduction Trehalose, a glass-forming bioprotectant disaccharide, has been demonstrated to possess significant potential within the food industry. It does not interact with reactive molecules such as amino groups from peptides and proteins, preventing the degradation and aggregation due to Maillard reactions. Objective This paper aims to review at the molecular level the effects of trehalose on the structural and dynamical properties of water and on protein to highlight the stabilization and conservation properties on food products. Results and Conclusions The experimental findings presented show that water molecules are arranged in presence of trehalose in a particular configuration which avoids ice formation, so limiting damage due to freezing and cooling. On the other hand, homologous disaccharides, and trehalose to a greater extent, slow down the dynamics of water with a significant influence on the biological activity. These results imply that trehalose has a greater ability to bind volatile substances and deliver superior bioprotective effectiveness. Furthermore trehalose is shown to be incapable of taking part in the denaturation process of lysozyme under thermal stress. [source] Modified Two-Layer Preservation Method (M-Kyoto/PFC) Improves Islet Yields in Islet IsolationAMERICAN JOURNAL OF TRANSPLANTATION, Issue 3 2006H. Noguchi Islet allotransplantation can achieve insulin independence in patients with type I diabetes. Recent reports show that the two-layer method (TLM), which employs oxygenated perfluorochemical (PFC) and UW solution, is superior to simple cold storage in UW for pancreas preservation in islet transplantation. However, UW solution has several disadvantages, including the inhibition of Liberase activity. In this study, we investigated the features of a new solution, designated M-Kyoto solution. M-Kyoto solution contains trehalose and ulinastatin as distinct components. Trehalose has a cytoprotective effect against stress, and ulinastatin inhibits trypsin. In porcine islet isolation, islet yield was significantly higher in the M-Kyoto/PFC group compared with the UW/PFC group. There was no significant difference in ATP content in the pancreas between the two groups, suggesting that different islet yields are not due to their differences as energy sources. Compared with UW solution, M-Kyoto solution significantly inhibited trypsin activity in the digestion step; moreover, M-Kyoto solution inhibited collagenase digestion less than UW solution. In conclusion, the advantages of M-Kyoto solution are trypsin inhibition and less collagenase inhibition. Based on these data, we now use M-Kyoto solution for clinical islet transplantation from nonheart-beating donor pancreata. [source] Metabolic profiling as a tool for understanding defense response of Taxus Cuspidata cells to shear stressBIOTECHNOLOGY PROGRESS, Issue 5 2009Pei-Pei Han Abstract To obtain a better understanding of responsive mechanism of plant cells in response to hydrodynamic mechanical stress, a metabolic profiling approach was used to profile metabolite changes of Taxus cuspidata cells under laminar shear stress. A total of 65 intracellular metabolites were identified and quantified, using gas chromatography coupled to time-of-flight mass spectrometry. Potential biomarkers were found by the principal component analysis as well as partial least squares combined with variable influence in the projection. Trehalose, sorbitol, ascorbate, sucrose, and gluconic acid were mainly responsible for the discrimination between shear stress induced cells and control cells. Further analysis by mapping measured metabolite concentrations onto the metabolic network revealed that shear stress imposed restrictions on primary metabolic pathways by inhibiting tricarboxylic acid cycle, glycolysis, and N metabolism. To adapt to the shear condition, cells responded by starting defensive programs. These defensive programs included coinduction of glycolysis and sucrose metabolism, accumulation of compatible solutes, and antioxidative strategy. A strategy of defense mechanisms at the level of metabolites for T. cuspidata cells when challenged with the shear stress was proposed. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source] Cross-correlated and conventional dipolar carbon-13 relaxation in methylene groups in small, symmetric moleculesCONCEPTS IN MAGNETIC RESONANCE, Issue 2 2007Leila Ghalebani Abstract A theory for dipolar cross-correlated relaxation processes in AMX or AX2 spin system, with special reference to 13C-methylene groups, is reviewed briefly. Simple experiments and protocols for measuring the transfer rates between the carbon-13 Zeeman order and the three-spin order, and for their analogues in the transverse plane, are discussed using a concentrated solution of the disaccharide trehalose as a model system. Experimental data sets consisting of conventional carbon-13 relaxation parameters (T1, T2, and NOE), along with the cross-correlated relaxation rates, are also presented for some small, rigid, polycyclic molecules. These data are interpreted using spectral density functions appropriate to spherical or symmetric tops reorienting according to small-step rotational diffusion model. The analysis results in a consistent picture of the auto- and cross-correlated spin relaxation processes. © 2007 Wiley Periodicals, Inc. Concepts Magn Reson Part A 30A: 100,115, 2007. [source] Universal and species-specific bacterial ,fungiphiles' in the mycospheres of different basidiomycetous fungiENVIRONMENTAL MICROBIOLOGY, Issue 2 2009J. A. Warmink Summary In previous work, several bacterial groups that show a response to fruiting bodies (the mycosphere) of the ectomycorrhizal fungus Laccaria proxima were identified. We here extend this work to a broader range of fungal fruiting bodies sampled at two occasions. PCR-DGGE analyses showed clear effects of the mycosphere of diverse fungi on the total bacterial and Pseudomonas communities in comparison with those in the corresponding bulk soil. The diversities of the Pseudomonas communities increased dramatically in most of the mycospheres tested, which contrasted with a decrease of the diversity of the total bacterial communities in these habitats. The data also indicated the existence of universal (i.e. Pseudomonas poae, P. lini, P. umsongensis, P. corrugata, P. antarctica and Rahnella aquatilis) as well as specific (i.e. P. viridiflava and candidatus Xiphinematobacter americani) fungiphiles, defined as bacteria adapted to the mycospheres of, respectively, three or more or just one fungal species. The selection of such fungiphiles was shown to be strongly related to their capacities to use particular carbonaceous compounds, as evidenced using principal components analyses of BIOLOG-based substrate utilization tests. The differentiating compounds, i.e. l -arabinose, l -leucine, m-inositol, m-arabitol, d -mannitol and d -trehalose, were tentatively linked to compounds known to occur in mycosphere exudates. [source] Adenylyl cyclase encoded by AC78C participates in sugar perception in Drosophila melanogasterEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 10 2008Kohei Ueno Abstract In gustatory receptor neurons (GRNs) in Drosophila melanogaster, Gr5a and one of the Gr64s encode sugar receptors with seven transmembrane domains. Previously, we have shown that the responses to various sugars are depressed in DGs, mutant flies (Ueno et al., 2006). Because DGs, is a homolog of Gs, we hypothesized that the sugar receptors are coupled to adenylyl cyclase (AC) in Drosophila. The aim of this study was to identify the AC that participates in sugar perception. Here, we found that an AC inhibitor, MDL-12330A, depressed the response in GRNs to trehalose as well as sucrose; that an AC gene, AC78C, was expressed in the sugar-sensitive GRNs; that RNAi against AC78C depressed the electrical response in GRNs to sucrose; and that the sugar response in GRNs, as well as sugar intake in a behavioral assay in an AC78C mutant, was depressed at low sugar concentrations. We conclude that AC78C, via cAMP, participates in the sugar-taste signaling pathway at the low concentration range. [source] Development of Ketoside-Type Analogues of Trehalose by Using ,-Stereoselective O-Glycosidation of KetoseEUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 22 2007Rie Namme Abstract The stereoselective synthesis of ketoside-type analogues of trehalose is described. O-Glycosidation of hept-2-ulopyranose with trimethylsilyl ,-pyranoside promoted by trimethylsilyl trifluoromethanesulfonate afforded ,-ketopyranosyl ,-aldopyranosides exclusively. ,-Ketopyranosyl ,-aldooyranosides and ,-ketopyranosyl ,-ketopyranosides were also synthesized in a similar manner. The benzyl protecting groups of the hydroxy moieties were removed by hydrogenolysis to afford fully deprotected trehalose analogues.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Enzymatic control of anhydrobiosis-related accumulation of trehalose in the sleeping chironomid, Polypedilum vanderplankiFEBS JOURNAL, Issue 20 2010Kanako Mitsumasu Larvae of an anhydrobiotic insect, Polypedilum vanderplanki, accumulate very large amounts of trehalose as a compatible solute on desiccation, but the molecular mechanisms underlying this accumulation are unclear. We therefore isolated the genes coding for trehalose metabolism enzymes, i.e. trehalose-6-phosphate synthase (TPS) and trehalose-6-phosphate phosphatase (TPP) for the synthesis step, and trehalase (TREH) for the degradation step. Although computational prediction indicated that the alternative splicing variants (PvTps,/,) obtained encoded probable functional motifs consisting of a typical consensus domain of TPS and a conserved sequence of TPP, PvTps, did not exert activity as TPP, but only as TPS. Instead, a distinct gene (PvTpp) obtained expressed TPP activity. Previous reports have suggested that insect TPS is, exceptionally, a bifunctional enzyme governing both TPS and TPP. In this article, we propose that TPS and TPP activities in insects can be attributed to discrete genes. The translated product of the TREH ortholog (PvTreh) certainly degraded trehalose to glucose. Trehalose was synthesized abundantly, consistent with increased activities of TPS and TPP and suppressed TREH activity. These results show that trehalose accumulation observed during anhydrobiosis induction in desiccating larvae can be attributed to the activation of the trehalose synthetic pathway and to the depression of trehalose hydrolysis. [source] Aspergillus nidulans,-galactosidase of glycoside hydrolase family 36 catalyses the formation of ,-galacto-oligosaccharides by transglycosylationFEBS JOURNAL, Issue 17 2010Hiroyuki Nakai The ,-galactosidase from Aspergillus nidulans (AglC) belongs to a phylogenetic cluster containing eukaryotic ,-galactosidases and ,-galacto-oligosaccharide synthases of glycoside hydrolase family 36 (GH36). The recombinant AglC, produced in high yield (0.65 g·L,1 culture) as His-tag fusion in Escherichia coli, catalysed efficient transglycosylation with ,-(1,6) regioselectivity from 40 mm 4-nitrophenol ,- d -galactopyranoside, melibiose or raffinose, resulting in a 37,74% yield of 4-nitrophenol ,- d -Galp -(1,6)- d -Galp, ,- d -Galp -(1,6)-,- d -Galp -(1,6)- d -Glcp and ,- d -Galp -(1,6)-,- d -Galp -(1,6)- d -Glcp -(,1,,2)- d -Fruf (stachyose), respectively. Furthermore, among 10 monosaccharide acceptor candidates (400 mm) and the donor 4-nitrophenol ,- d -galactopyranoside (40 mm), ,-(1,6) linked galactodisaccharides were also obtained with galactose, glucose and mannose in high yields of 39,58%. AglC did not transglycosylate monosaccharides without the 6-hydroxymethyl group, i.e. xylose, l -arabinose, l -fucose and l -rhamnose, or with axial 3-OH, i.e. gulose, allose, altrose and l -rhamnose. Structural modelling using Thermotoga maritima GH36 ,-galactosidase as the template and superimposition of melibiose from the complex with human GH27 ,-galactosidase supported that recognition at subsite +1 in AglC presumably requires a hydrogen bond between 3-OH and Trp358 and a hydrophobic environment around the C-6 hydroxymethyl group. In addition, successful transglycosylation of eight of 10 disaccharides (400 mm), except xylobiose and arabinobiose, indicated broad specificity for interaction with the +2 subsite. AglC thus transferred ,-galactosyl to 6-OH of the terminal residue in the ,-linked melibiose, maltose, trehalose, sucrose and turanose in 6,46% yield and the ,-linked lactose, lactulose and cellobiose in 28,38% yield. The product structures were identified using NMR and ESI-MS and five of the 13 identified products were novel, i.e. ,- d -Galp -(1,6)- d -Manp; ,- d -Galp -(1,6)-,- d -Glcp -(1,4)- d -Glcp; ,- d -Galp -(1,6)-,- d -Galp -(1,4)- d -Fruf; ,- d -Galp -(1,6)- d -Glcp -(,1,,1)- d -Glcp; and ,- d -Galp -(1,6)-,- d -Glcp -(1,3)- d -Fruf. [source] The phosphate site of trehalose phosphorylase from Schizophyllum commune probed by site-directed mutagenesis and chemical rescue studiesFEBS JOURNAL, Issue 5 2008Christiane Goedl Schizophyllum commune,,,-trehalose phosphorylase utilizes a glycosyltransferase-like catalytic mechanism to convert its disaccharide substrate into ,- d -glucose 1-phosphate and ,- d -glucose. Recruitment of phosphate by the free enzyme induces ,,,-trehalose binding recognition and promotes the catalytic steps. Like the structurally related glycogen phosphorylase and other retaining glycosyltransferases of fold family GT-B, the trehalose phosphorylase contains an Arg507-XXXX-Lys512 consensus motif (where X is any amino acid) comprising key residues of its putative phosphate-binding sub-site. Loss of wild-type catalytic efficiency for reaction with phosphate (kcat/Km = 21 000 m,1·s,1) was dramatic (,107 -fold) in purified Arg507,Ala (R507A) and Lys512,Ala (K512A) enzymes, reflecting a corresponding change of comparable magnitude in kcat (Arg507) and Km (Lys512). External amine and guanidine derivatives selectively enhanced the activity of the K512A mutant and the R507A mutant respectively. Analysis of the pH dependence of chemical rescue of the K512A mutant by propargylamine suggested that unprotonated amine in combination with H2PO4,, the protonic form of phosphate presumably utilized in enzymatic catalysis, caused restoration of activity. Transition state-like inhibition of the wild-type enzyme A by vanadate in combination with ,,,-trehalose (Ki = 0.4 ,m) was completely disrupted in the R507A mutant but only weakened in the K512A mutant (Ki = 300 ,m). Phosphate (50 mm) enhanced the basal hydrolase activity of the K512A mutant toward ,,,-trehalose by 60% but caused its total suppression in wild-type and R507A enzymes. The results portray differential roles for the side chains of Lys512 and Arg507 in trehalose phosphorylase catalysis, reactant state binding of phosphate and selective stabilization of the transition state respectively. [source] Biochemical characterization of rice trehalose-6-phosphate phosphatases supports distinctive functions of these plant enzymesFEBS JOURNAL, Issue 5 2007Shuhei Shima Substantial levels of trehalose accumulate in bacteria, fungi, and invertebrates, where it serves as a storage carbohydrate or as a protectant against environmental stresses. In higher plants, trehalose is detected at fairly low levels; therefore, a regulatory or signaling function has been proposed for this molecule. In many organisms, trehalose-6-phosphate phosphatase is the enzyme governing the final step of trehalose biosynthesis. Here we report that OsTPP1 and OsTPP2 are the two major trehalose-6-phosphate phosphatase genes expressed in vegetative tissues of rice. Similar to results obtained from our previous OsTPP1 study, complementation analysis of a yeast trehalose-6-phosphate phosphatase mutant and activity measurement of the recombinant protein demonstrated that OsTPP2 encodes a functional trehalose-6-phosphate phosphatase enzyme. OsTPP2 expression is transiently induced in response to chilling and other abiotic stresses. Enzymatic characterization of recombinant OsTPP1 and OsTPP2 revealed stringent substrate specificity for trehalose 6-phosphate and about 10 times lower Km values for trehalose 6-phosphate as compared with trehalose-6-phosphate phosphatase enzymes from microorganisms. OsTPP1 and OsTPP2 also clearly contrasted with microbial enzymes, in that they are generally unstable, almost completely losing activity when subjected to heat treatment at 50 °C for 4 min. These characteristics of rice trehalose-6-phosphate phosphatase enzymes are consistent with very low cellular substrate concentration and tightly regulated gene expression. These data also support a plant-specific function of trehalose biosynthesis in response to environmental stresses. [source] Physiological and morphological responses of the soil bacterium Rhodococcus opacus strain PD630 to water stressFEMS MICROBIOLOGY ECOLOGY, Issue 2 2004Héctor M. Alvarez Abstract Rhodococcus opacus PD630 was investigated for physiological and morphological changes under water stress challenge. Gluconate- and hexadecane-grown cells were extremely resistant to these conditions, and survival accounted for up to 300 and 400 days; respectively, when they were subjected to slow air-drying. Results of this study suggest that strain PD630 has specific mechanisms to withstand water stress. Water-stressed cells were sensitive to the application of ethanol, high temperatures and oxidative stress, whereas they exhibited cross-protection solely against osmotic stress during the first hours of application. Results indicate that the resistance programme for water stress in R. opacus PD630 includes the following physiological and morphological changes, among others: (1) energetic adjustments with drastic reduction of the metabolic activity (,39% decrease during the first 24 h and about 90% after 190 days under dehydration), (2) endogenous metabolism using intracellular triacylglycerols for generating energy and precursors, (3) biosynthesis of different osmolytes such as trehalose, ectoine and hydroxyectoine, which may achieve a water balance through osmotic adjustment and may explain the overlap between water and osmotic stress, (4) adjustments of the cell-wall through the turnover of mycolic acid species, as preliminary experiments revealed no evident changes in the thickness of the cell envelope, (5) formation of short fragmenting-cells as probable resistance forms, (6) production of an extracellular slime covering the surface of colonies, which probably regulates internal and external c anges in water potential, and (7) formation of compact masses of cells. This contributes to understanding the water stress resistance processes in the soil bacterium R. opacus PD630. [source] Adhesion and development of the root rot fungus (Heterobasidion annosum) on conifer tissues: effects of spore and host surface constituentsFEMS MICROBIOLOGY ECOLOGY, Issue 2 2000Frederick O Asiegbu Abstract The objective of this study was to correlate the occurrence of particular root and woody stump surface components with the ability of spores of the root rot fungus (Heterobasidion annosum) to adhere, germinate and establish on conifer tissues. With the aid of high performance liquid chromatography, several sugars (pinitol, xylitol, dulcitol, mannitol, D -glucose, mannose, fructose) were detected on both stump and fine root surfaces of Scots pine and Norway spruce. Of all the sugars observed, xylose and arabinose were poorly utilized for initiation of germ tube growth whereas spore germination was enhanced in the presence of D -glucose, mannose or fructose. Oxidation of these sugars by pretreatment of wood discs or roots with periodic acid abolished the ability of the spores to germinate. Non-sugar components such as long chain fatty acids on spores and root surfaces as detected with nuclear magnetic resonance were found to have a significant influence on adhesion and initiation of germ tube development. Removal of these aliphatic compounds from the root surface increased spore germination by 2-fold, whereas similar treatment on spores led to a 5-fold decrease in adhesiveness to root material. In vitro studies revealed that the di-ethyl ether extract from the roots had no long term adverse effect on spore germination which suggests that the fungus may possess the capability to detoxify this substance. Similarly, adhesion of spores was affected by low and freezing temperatures. The role of significant levels of mannitol and trehalose accumulated in spores and hyphae of the fungi on viability, survival and tolerance to adverse conditions such as oxidative stress, freezing and desiccation are discussed. [source] Adaptive tolerance to oxidative stress and the induction of antioxidant enzymatic activities in Candida albicans are independent of the Hog1 and Cap1-mediated pathwaysFEMS YEAST RESEARCH, Issue 6 2010Pilar Gónzalez-Párraga Abstract In the pathogenic yeast Candida albicans, the MAP-kinase Hog1 mediates an essential protective role against oxidative stress, a feature shared with the transcription factor Cap1. We analysed the adaptive oxidative response of strains with both elements altered. Pretreatment with gentle doses of oxidants or thermal upshifts (28,37 and 37,42 °C) improved survival in the face of high concentrations of oxidants (50 mM H2O2 or 40 mM menadione), pointing to a functional cross-protective mechanism in the mutants. The oxidative challenge promoted a marked intracellular synthesis of trehalose, although hog1 (but not cap1) cells always displayed high basal trehalose levels. Hydrogen peroxide (H2O2) induced mRNA expression of the trehalose biosynthetic genes (TPS1 and TPS2) in the tested strains. Furthermore, oxidative stress also triggered a differential activation of various antioxidant activities, whose intensity was greater after HOG1 and CAP1 deletion. The pattern of activity was dependent on the oxidant dosage applied: low concentrations of H2O2 (0.5,5 mM) clearly induced catalase and glutathione reductase (GR), whereas drastic H2O2 exposure (50 mM) increased Mn-superoxide dismutase (SOD) isozyme-mediated SOD activity. These results firmly support the existence in C. albicans of both Hog1- and Cap1-independent mechanisms against oxidative stress. [source] Anaerobic homolactate fermentation with Saccharomyces cerevisiae results in depletion of ATP and impaired metabolic activityFEMS YEAST RESEARCH, Issue 3 2009Derek A. Abbott Abstract Conversion of glucose to lactic acid is stoichiometrically equivalent to ethanol formation with respect to ATP formation from substrate-level phosphorylation, redox equivalents and product yield. However, anaerobic growth cannot be sustained in homolactate fermenting Saccharomyces cerevisiae. ATP-dependent export of the lactate anion and/or proton, resulting in net zero ATP formation, is suspected as the underlying cause. In an effort to understand the mechanisms behind the decreased lactic acid production rate in anaerobic homolactate cultures of S. cerevisiae, aerobic carbon-limited chemostats were performed and subjected to anaerobic perturbations in the presence of high glucose concentrations. Intracellular measurements of adenosine phosphates confirmed ATP depletion and decreased energy charge immediately upon anaerobicity. Unexpectedly, readily available sources of carbon and energy, trehalose and glycogen, were not activated in homolactate strains as they were in reference strains that produce ethanol. Finally, the anticipated increase in maximal velocity (Vmax) of glycolytic enzymes was not observed in homolactate fermentation suggesting the absence of protein synthesis that may be attributed to decreased energy availability. Essentially, anaerobic homolactate fermentation results in energy depletion, which, in turn, hinders protein synthesis, central carbon metabolism and subsequent energy generation. [source] Role of reserve carbohydrates in the growth dynamics of Saccharomyces cerevisiae,FEMS YEAST RESEARCH, Issue 8 2004Vincent Guillou Abstract The purpose of this study was to explore the role of glycogen and trehalose in the ability of Saccharomyces cerevisiae to respond to a sudden rise of the carbon flux. To this end, aerobic glucose-limited continuous cultures were challenged with a sudden increase of the dilution rate from 0.05 to 0.15 h,1. Under this condition, a rapid mobilization of glycogen and trehalose was observed which coincided with a transient burst of budding and a decrease of cell biomass. Experiments carried out with mutants defective in storage carbohydrates indicated a predominant role of glycogen in the adaptation to this perturbation. However, the real importance of trehalose in this response was veiled by the unexpected phenotypes harboured by the tps1 mutant, chosen for its inability to synthesize trehalose. First, the biomass yield of this mutant was 25% lower than that of the isogenic wild-type strain at dilution rate of 0.05 h,1, and this difference was annulled when cultures were run at a higher dilution rate of 0.15 h,1. Second, the tps1 mutant was more effective to sustain the dilution rate shift-up, apparently because it had a faster glycolytic rate and an apparent higher capacity to consume glucose with oxidative phosphorylation than the wild type. Consequently, a tps1gsy1gsy2 mutant was able to adapt to the dilution rate shift-up after a long delay, likely because the detrimental effects from the absence of glycogen was compensated for by the tps1 mutation. Third, a glg1,glg2, strain, defective in glycogen synthesis because of the lack of the glycogen initiation protein, recovered glycogen accumulation upon further deletion of TPS1. This recovery, however, required glycogen synthase. Finally, we demonstrated that the rapid breakdown of reserve carbohydrates triggered by the shift-up is merely due to changes in the concentrations of hexose-6-phosphate and UDPglucose, which are the main metabolic effectors of the rate-limiting enzymes of glycogen and trehalose pathways. [source] The importance of a functional trehalose biosynthetic pathway for the life of yeasts and fungiFEMS YEAST RESEARCH, Issue 4-5 2004Carlos Gancedo Abstract The view of the role of trehalose in yeast has changed in the last few years. For a long time considered a reserve carbohydrate, it gained new importance when its function in the acquisition of thermotolerance was demonstrated. More recently the cellular processes in which the trehalose biosynthetic pathway has been implicated range from the control of glycolysis to sporulation and infectivity by certain fungal pathogens. There is now enough experimental evidence to conclude that trehalose 6-phosphate, an intermediate of trehalose biosynthesis, is an important metabolic regulator in such different organisms as yeasts or plants. Its inhibition of hexokinase plays a key role in the control of the glycolytic flux in Saccharomyces cerevisiae but other, likely important, sites of action are still unknown. We present examples of the phenotypes produced by mutations in the two steps of the trehalose biosynthetic pathway in different yeasts and fungi, and whenever possible examine the molecular explanations advanced to interpret them. [source] Cloning and characterization of genes encoding trehalose-6-phosphate synthase (TPS1) and trehalose-6-phosphate phosphatase (TPS2) from Zygosaccharomyces rouxiiFEMS YEAST RESEARCH, Issue 4 2003Hawk-Bin Kwon Abstract In many organisms, trehalose protects against several environmental stresses, such as heat, desiccation, and salt, probably by stabilizing protein structures and lipid membranes. Trehalose synthesis in yeast is mediated by a complex of trehalose-6-phosphate synthase (TPS1) and trehalose-6-phosphate phosphatase (TPS2). In this study, genes encoding TPS1 and TPS2 were isolated from Zygosaccharomyces rouxii (designated ZrTPS1 and ZrTPS2, respectively). They were functionally identified by their complementation of the tps1 and tps2 yeast deletion mutants, which are unable to grow on glucose medium and with heat, respectively. Full-length ZrTPS1 cDNA is composed of 1476 nucleotides encoding a protein of 492 amino acids with a molecular mass of 56 kDa. ZrTPS2 cDNA consists of 2843 nucleotides with an open reading frame of 2700 bp, which encodes a polypeptide of 900 amino acids with a molecular mass of 104 kDa. The amino acid sequence encoded by ZrTPS1 has relatively high homology with TPS1 of Saccharomyces cerevisiae and Schizosaccharomyces pombe, compared with TPS2. Western blot analysis showed that the antibody against S. cerevisiae TPS1 recognizes ZrTPS1. Under normal growth conditions, ZrTPS1 and ZrTPS2 were highly and constitutively expressed, unlike S. cerevisiae TPS1 and TPS2. Salt stress and heat stress reduced the expression of the ZrTPS1 and ZrTPS2 genes, respectively. [source] Dehydration tolerance and water vapour absorption in two species of soil-dwelling Collembola by accumulation of sugars and polyolsFUNCTIONAL ECOLOGY, Issue 5 2001M. Holmstrup Summary 1,Physiological and biochemical responses to desiccation were studied in two species of hygrophilic Collembola, Folsomia fimetaria L. and Protaphorura armata Tullberg. The temporal changes in water content, body fluid osmotic pressure, and accumulation of sugars and polyols during desiccation stress, equivalent to severe soil drought are reported. 2,Both species initially lost about 50% of their initial content of osmotically active water. Within 7 days F. fimetaria had re-established their preliminary water content by absorbing water vapour from the atmosphere. Protaphorura armata also significantly increased their water content over the following 10 days. 3,Both species were able to resume their hyperosmotic status relative to the desiccating environment they were placed in. The accumulation of myo-inositol and two other unidentified compounds (F. fimetaria) and trehalose (P. armata) made an important contribution to this phenomenon. 4,These results suggest that water vapour absorption by accumulation of sugars and polyols may be widespread in soil-dwelling Collembola. [source] Effects of physiological isotonic cryoprotectants on living cells during the freezing,thawing process and effects of their uptake by electroporation: Sp2 cells in alginate,trehalose solutionsHEAT TRANSFER - ASIAN RESEARCH (FORMERLY HEAT TRANSFER-JAPANESE RESEARCH), Issue 6 2003Ryo Shirakashi Abstract In the present study, effects of isotonic alginate and trehalose solutions on electroporated and unpulsed control murine myeloma cells (Sp2 line) during the freezing,thawing process were evaluated. To investigate the effects of these potential cryoprotectants, microscopic examination of cell suspensions in alginate,NaCl, alginate,trehalose, or trehalose media was performed during freezing,thawing at various cooling/warming rates (e.g., ,1, ,5, and ,90 °C/min). It was found that, even at the lowest cooling rate, extracellular ice grains did not mechanically suppress cells in the trehalose-substituted medium. Moreover, at all cooling rates, intracellular ice nucleation seldom occurred in the presence of extracellular trehalose. © 2003 Wiley Periodicals, Inc. Heat Trans Asian Res, 32(6): 511,523, 2003; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/htj.10105 [source] | |||||||||||||||||||||||||||||||||||||||||||