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PubMed1979

Uptake of trehalose by Saccharomyces cerevisiae.

Trehalose, a storage sugar of baker's yeast, is known not to be metabolized when added to a cell suspension in water or a growth medium and to support growth only after a lag of about 10 h. However, it was transported into cells by at least two transport systems, the uptake being active, with a pH optimum at 5.5. There was no stoicheiometry with the shift of protons into cells observed at high trehalose concentrations. Trehalose remained intact in cells and was not appreciably lost to a trehalose-free medium. The uptake systems were present directly after growth on glucose, then decayed with a half-life of about 25 min but could be reactivated by aerobic incubation with trehalose, maltose, alpha-methyl-D-glucoside, glucose or ethanol. The uptake systems thus induced were different as revealed by competition experiments. At least one of the systems for trehalose uptake showed cooperative kinetics. Comparative anaysis with other disaccharides indicated the existence in Saccharomyces cerevisiae, after induction with trehalose, of at least four systems for the uptake of alpha-methyl-D-glucoside, four systems for maltose, together with the two for trehalose, variously shared by the sugars, the total of alpha-glucoside-transporting systems being five.

Antimetabolites↗
PubMed1996

The orlA gene from Aspergillus nidulans encodes a trehalose-6-phosphate phosphatase necessary for normal growth and chitin synthesis at elevated temperatures.

A cosmid carrying the orlA gene from Aspergillus nidulans was identified by complementation of an orlA1 mutant strain with DNA from the pKBY2 cosmid library. An orlA1 complementing fragment from the cosmid was sequenced. orlA encodes a predicted polypeptide of 227 amino acids (26360 Da) that is homologous to a 211-amino-acid domain from the polypeptide encoded by the Saccharomyces cerevisiae TPS2 gene and to almost the entire Escherichia coli otsB-encoded polypeptide. TPS2 and otsB each specify a trehalose-6-phosphate phosphatase, an enzyme that is necessary for trehalose synthesis. orlA disruptants accumulate trehalose-6-phosphate and have reduced trehalose-6-phosphatate phosphatase levels, indicating that the gene encodes a trehalose-6-phosphatate phosphatase. Disruptants have a nearly-wild-type morphology at 32 degrees C. When germinated at 42 degrees C, the conidia and hyphae from disruptants are chitin deficient, swell excessively, and lyse. The lysis is almost completely remedied by osmotic stabilizers and is partially remedied by N-acetylglucosamine (GlcNAc). The activity of glutamine:fructose-6-phosphate amido-transferase (GFAT), the first enzyme unique to aminosugar synthesis, is reduced and is labile in orlA disruption strains. The findings are consistent with the hypothesis that trehalose-6-phosphate reduces the temperature stability of GFAT and other enzymes of chitin metabolism at elevated temperatures. The results extend to filamentous organisms the observation that mutations in fungal trehalose synthesis are highly pleiotropic and affect aspects of carbohydrate metabolism that are not directly related to trehalose synthesis.

Amino Acid Sequence↗
PubMed1996

Kinetics and energetics of trehalose transport in Saccharomyces cerevisiae.

Cells of Saccharomyces cerevisiae are able to transport trehalose against a concentration gradient, without efflux or counterflow of the labeled substrate. Uptake was inhibited by uncouplers, acetic acid, and organic mercury compounds. The addition of trehalose resulted in alkalinization of the medium. The ratio of H+ depletion to trehalose uptake by yeast cells was approximately 1:1, which indicates the existence of a trehalose-H+ symporter in these cells. The optimum pH for this active H+-trehalose symport was 5.0, and both the Km and the Vmax were negatively affected by increasing or decreasing the extracellular pH from its optimum value. Kinetic studies showed the existence of at least two different trehalose transport activities in yeast cells: a high-affinity H+-trehalose symporter (Km = 4 mM), and a low-affinity transport activity (Km > 100 mM) that could be a facilitated diffusion process. The high-affinity H+-trehalose symporter was repressed by glucose, whereas the low-affinity uptake was constitutively expressed in S. cerevisiae.

Biological Transport, Active↗
PubMed2006

Intracellular trehalose is neither necessary nor sufficient for desiccation tolerance in yeast.

Trehalose is thought to be important for desiccation tolerance in a number of organisms, including Saccharomyces cerevisiae, but there is limited in vivo evidence to support this hypothesis. In wild-type yeast, the degree of desiccation tolerance has been shown previously to increase in cultures after diauxic shift and also in exponential-phase cultures after exposure to heat stress. Under both these conditions, increased survival of desiccation correlates with elevated intracellular trehalose concentrations. Our data confirm these findings, but we have tested the apparent importance of trehalose using mutant strains with a deleted trehalose-6-phosphate synthase gene (tps1Delta). Although tps1Delta strains do not produce trehalose, they are nevertheless capable of desiccation tolerance, and the degree of tolerance also increases after diauxic shift or heat stress, albeit slightly less than in the wild type. Conversely, when wild-type yeast is subjected to osmotic stress, mid-exponential-phase cultures produce high concentrations of intracellular trehalose but show little improvement in desiccation tolerance. These results show that there is no consistent relationship between intracellular trehalose levels and desiccation tolerance in S. cerevisiae. Trehalose seems to be neither necessary nor sufficient for, although in some strains might quantitatively improve, survival of desiccation, suggesting that other adaptations are more important.

Cytoplasm↗
PubMed1989

Trehalose accumulation in vegetative cells and spores of Myxococcus xanthus.

The disaccharide trehalose is found in the spores and cysts of a variety of organisms. We analyzed developing cells of Myxococcus xanthus for trehalose accumulation. Vegetative cells grown in media with low osmotic strengths contained less than 5 micrograms of trehalose per mg of protein. Spores formed in fruiting bodies accumulated up to 1,100 micrograms of trehalose per mg of protein. Spores formed in liquid culture following the addition of glycerol contained up to 300 micrograms of trehalose per mg of protein. The trehalose contents of both spore types decreased rapidly during the early stages of germination. Trehalase activity was not detected in extracts of dormant or germinating spores. Trehalose accumulation in M. xanthus was also associated with elevated osmotic strength. Vegetative cells accumulated up to 214 micrograms of trehalose per mg of protein when grown in media containing elevated levels of solutes.

Disaccharides↗
PubMed1991

Synthesis, accumulation, and excretion of trehalose in osmotically stressed Escherichia coli K-12 strains: influence of amber suppressors and function of the periplasmic trehalase.

It has been reported previously that Escherichia coli K-12 carries an amber mutation that prevents osmotic stress-dependent accumulation of trehalose (M. L. Rod, K. Y. Alam, P. R. Cunningham, and D. P. Clark, J. Bacteriol. 170:3601-3610, 1988). We report that E. coli K-12 and W1485 (sup0) accumulated trehalose but that they required a higher osmotic strength in the growth medium than that required by their sup+ derivatives. Furthermore, the sup+ derivatives displayed both strongly increased trehalose-6-phosphate synthase activity and expression of otsA-lacZ and otsB-lacZ operon fusions compared with their parental strains. It is suggested that the amber mutation in question may be in a gene system encoding a transcriptional activator of the ots genes which govern the synthase. The much-used sup0 strain MC4100 behaved like the sup+ derivatives of W1485 with respect to trehalose synthesis. treA mutants with a defective periplasmic trehalase accumulated trehalose extracellularly under osmotic stress. The amount of trehalose excreted correlated with their synthase activity. Strains with an intact trehalase did not display extracellular trehalose accumulation. Thus, stressed E. coli cells regulate the cytoplasmic level of trehalose by a futile cycle involving overproduction, excretion, and degradation to glucose, which is reutilized.

Escherichia coli↗
PubMed1994

Trehalose-6-P synthase is dispensable for growth on glucose but not for spore germination in Schizosaccharomyces pombe.

Trehalose-6-P inhibits hexokinases in Saccharomyces cerevisiae (M. A. Blázquez, R. Lagunas, C. Gancedo, and J. M. Gancedo, FEBS Lett. 329:51-54, 1993), and disruption of the TPS1 gene (formerly named CIF1 or FDP1) encoding trehalose-6-P synthase prevents growth in glucose. We have found that the hexokinase from Schizosaccharomyces pombe is not inhibited by trehalose-6-P even at a concentration of 3 mM. The highest internal concentration of trehalose-6-P that we measured in S. pombe was 0.75 mM after heat shock. We have isolated from S. pombe the tps1+ gene, which is homologous to the Saccharomyces cerevisiae TPS1 gene. The DNA sequence from tps1+ predicts a protein of 479 amino acids with 65% identity with the protein of S. cerevisiae. The tps1+ gene expressed from its own promoter could complement the lack of trehalose-6-P synthase in S. cerevisiae tps1 mutants. The TPS1 gene from S. cerevisiae could also restore trehalose synthesis in S. pombe tps1 mutants. A chromosomal disruption of the tps1+ gene in S. pombe did not have a noticeable effect on growth in glucose, in contrast with the disruption of TPS1 in S. cerevisiae. However, the disruption prevented germination of spores carrying it. The level of an RNA hybridizing with an internal probe of the tps1+ gene reached a maximum after 20 min of heat shock treatment. The results presented support the idea that trehalose-6-P plays a role in the control of glycolysis in S. cerevisiae but not in S. pombe and show that the trehalose pathway has different roles in the two yeast species.

Amino Acid Sequence↗
PubMed2001

Role of trehalose in growth at high temperature of Salmonella enterica serovar Typhimurium.

Moderate osmolality can stimulate bacterial growth at temperatures near the upper limit for growth. We investigated the mechanism by which high osmolality enhances the thermotolerance of Salmonella enterica serovar Typhimurium, by isolating bacteriophage MudI1734-induced insertion mutations that blocked the growth-stimulatory effect of 0.2 M NaCl at 45 degrees C. One of these mutations proved to be in the seqA gene (a regulator of initiation of DNA synthesis). Because this gene is cotranscribed with pgm (which encodes phosphoglucomutase), it is likely to be polar on the expression of the pgm gene. Pgm catalyzes the conversion of glucose-6-phosphate to glucose-1-phosphate during growth on glucose, and therefore loss of Pgm results in a deficiency in a variety of cellular constituents derived from glucose-1-phosphate, including trehalose. To test the possibility that the growth defect of the seqA::MudI1734 mutant at high temperature in medium of high osmolality is due to the block in trehalose synthesis, we determined the effect of an otsA mutation, which inactivates the first step of the trehalose biosynthetic pathway. The otsA mutation caused a growth defect at 45 degrees C in minimal medium containing 0.2 M NaCl that was similar to that caused by the pgm mutation, but otsA did not affect growth rate in this medium at 37 degrees C. These results suggest that the growth defect of the seqA-pgm mutant at high temperature could be a consequence of the block in trehalose synthesis. We found that, in addition to the well-known osmotic control, there is a temperature-dependent control of trehalose synthesis such that, in medium containing 0.2 M NaCl, cells grown at 45 degrees C had a fivefold higher trehalose pool size than cells grown at 30 degrees C. Our observations that trehalose accumulation is thermoregulated and that mutations that block trehalose synthesis cause a growth defect at high temperature in media of high osmolality suggested that this disaccharide is crucial for growth at high temperature either for turgor maintenance or for protein stabilization.

Bacterial Outer Membrane Proteins↗
PubMed2002

Redundancy in periplasmic binding protein-dependent transport systems for trehalose, sucrose, and maltose in Sinorhizobium meliloti.

We have identified a cluster of six genes involved in trehalose transport and utilization (thu) in Sinorhizobium meliloti. Four of these genes, thuE, -F, -G, and -K, were found to encode components of a binding protein-dependent trehalose/maltose/sucrose ABC transporter. Their deduced gene products comprise a trehalose/maltose-binding protein (ThuE), two integral membrane proteins (ThuF and ThuG), and an ATP-binding protein (ThuK). In addition, a putative regulatory protein (ThuR) was found divergently transcribed from the thuEFGK operon. When the thuE locus was inactivated by gene replacement, the resulting S. meliloti strain was impaired in its ability to grow on trehalose, and a significant retardation in growth was seen on maltose as well. The wild type and the thuE mutant were indistinguishable for growth on glucose and sucrose. This suggested a possible overlap in function of the thuEFGK operon with the aglEFGAK operon, which was identified as a binding protein-dependent ATP-binding transport system for sucrose, maltose, and trehalose. The K(m)s for trehalose transport were 8 +/- 1 nM and 55 +/- 5 nM in the uninduced and induced cultures, respectively. Transport and growth experiments using mutants impaired in either or both of these transport systems show that these systems form the major transport systems for trehalose, maltose, and sucrose. By using a thuE'-lacZ fusion, we show that thuE is induced only by trehalose and not by cellobiose, glucose, maltopentaose, maltose, mannitol, or sucrose, suggesting that the thuEFGK system is primarily targeted toward trehalose. The aglEFGAK operon, on the other hand, is induced primarily by sucrose and to a lesser extent by trehalose. Tests for root colonization, nodulation, and nitrogen fixation suggest that uptake of disaccharides can be critical for colonization of alfalfa roots but is not important for nodulation and nitrogen fixation per se.

ATP-Binding Cassette Transporters↗
PubMed2006

Insights on the evolution of trehalose biosynthesis.

BACKGROUND: The compatible solute trehalose is a non-reducing disaccharide, which accumulates upon heat, cold or osmotic stress. It was commonly accepted that trehalose is only present in extremophiles or cryptobiotic organisms. However, in recent years it has been shown that although higher plants do not accumulate trehalose at significant levels they have actively transcribed genes encoding the corresponding biosynthetic enzymes. RESULTS: In this study we show that trehalose biosynthesis ability is present in eubacteria, archaea, plants, fungi and animals. In bacteria there are five different biosynthetic routes, whereas in fungi, plants and animals there is only one. We present phylogenetic analyses of the trehalose-6-phosphate synthase (TPS) and trehalose-phosphatase (TPP) domains and show that there is a close evolutionary relationship between these domains in proteins from diverse organisms. In bacteria TPS and TPP genes are clustered, whereas in eukaryotes these domains are fused in a single protein. CONCLUSION: We have demonstrated that trehalose biosynthesis pathways are widely distributed in nature. Interestingly, several eubacterial species have multiple pathways, while eukaryotes have only the TPS/TPP pathway. Vertebrates lack trehalose biosynthetic capacity but can catabolise it. TPS and TPP domains have evolved mainly in parallel and it is likely that they have experienced several instances of gene duplication and lateral gene transfer.

Animals↗
PubMedDate not supplied

Uncoupling by trehalose of Ca2+ transport and ATP hydrolysis by the plasma membrane (Ca2+ + Mg2+) ATPase of kidney tubules.

Trehalose, the disaccharide of glucose, inhibits both initial rate and maximal capacity of ATP-dependent Ca2+ transport in inside-out vesicles of basolateral membrane from kidney proximal tubules. This inhibition (I0.5 = 60 mM) cannot be attributed to an increase in Ca2+ permeability, since the rate of EGTA-stimulated Ca2+ efflux from preloaded vesicles is not modified by trehalose. In the presence of 600 mM trehalose, Ca2+ uptake was almost completely inhibited, but the Ca(2+)-stimulated ATPase activity was unaffected; thus trehalose uncouples the Ca2+ transport from the ATPase activity. The Ca2+ transport inhibition by trehalose is reversible, since the inhibition disappeared when the vesicles were pre-incubated with 600 mM trehalose and then diluted in reaction medium to measure Ca2+ accumulation. Other mono- and disaccharides such as glucose, fructose, galactose, sucrose, maltose and lactose were tested but were not so effective as trehalose. The uncoupling of Ca2+ transport from hydrolysis can be explained by an interaction of trehalose with the phospholipid environment of the enzyme that induces conformational changes in specific domains of the enzyme so as to impair the coupling process.

Adenosine Triphosphate↗
PubMedDate not supplied

Protective role of trehalose in thermal denaturation of yeast pyrophosphatase.

Trehalose, a disaccharide of glucose, is accumulated in yeast cytosol when this organism is submitted to a stress condition. Recently it was shown that the level of trehalose increase up to 15 times when yeast cells are submitted to heat shock (De Virgilio et al., 1991). In this report we give evidence how trehalose may play an important role on the stress-survival of yeasts when submitted to a heat shock. We show that 1.5 M trehalose increases 13-fold the half-time for thermal inactivation (t0.5) of yeast cytosolic pyrophosphatase at 50 degrees C. This thermal protection conferred by trehalose is dose-dependent, after 10 min at 50 degrees C, a condition which inactivated pyrophosphatase, the presence of 2 M trehalose preserves 95% of total activity. Other carbohydrates were tested but were not so effective as trehalose. The presence of trehalose at high concentrations in the reaction medium at 35 degrees C inhibits pyrophosphatase activity. This inhibition is less effective at 50 degrees C suggesting that under this condition the enzyme is temperature-protected and active.

Disaccharides↗
PubMedDate not supplied

Effects of trehalose and ethanol on yeast cytosolic pyrophosphatase.

Trehalose has been described to protect several enzymes against destabilizing conditions. This sugar is naturally accumulated by yeast as a stress protectant. A common stress condition that yeast is normally submitted is the presence of ethanol, the by-product of fermentation process of several yeast. In this paper we show the effects of trehalose and ethanol, alone or together, on yeast pyrophosphatase, and the effects of these compounds on inhibition and unfolding of pyrophosphatase promoted by urea. We show that both trehalose and ethanol inhibit pyrophosphatase in a dose-dependent manner, and that the presence of ethanol does not modify the inhibition promoted by trehalose as well as the presence of trehalose does not modify the inhibition promoted by ethanol. The effects of trehalose on pyrophosphatase are completely reversible, but the inhibition caused by ethanol was only partially reversible. Incubation of pyrophosphatase with 10% (v/v) ethanol promoted an inhibition of 15%, and the control activity was completely recovered after removal of ethanol. On the other hand, when pyrophosphatase was incubated with 20% (v/v) ethanol an inhibition of 40% of the control activity was observed which persisted after removal of ethanol. Ethanol also potentiates the inhibition of pyrophosphatase promoted by urea, and contributes for an irreversible inactivation and unfolding of pyrophosphatase in the presence of urea. Trehalose, that protects this enzyme against the inhibition and unfolding promoted by the chaotropic compound urea, was inefficient to protect against the effects of ethanol. Trehalose was also efficient to prevent an irreversible inactivation induced by urea.

Ethanol↗
PubMed1998

Transitory laxative threshold of trehalose and lactulose in healthy women.

The transitory laxative threshold of a partially digestible disaccharide, trehalose, and an undigestible disaccharide, lactulose, was estimated by the dose-response relation between the test substance and the prevalence of diarrhea in 20 healthy female subjects. The subjects ingested several indicated amounts of trehalose or lactulose once daily 2 to 3 h after a meal. The intake of the test substance was stopped at the dose level that caused diarrhea or when the dose reached the maximal level. A record of physical conditions, gastrointestinal symptoms, and fecal conditions was made by all subjects before and after each ingestion of the test substance. Half the subjects experienced no diarrhea even with the ingestion of the maximal dose level (60 g) of trehalose in this study, and the ingestion of up to 40 g of lactulose caused diarrhea in 75% of all subjects. Abdominal symptoms such as flatus, distension, and borborygmus appeared at high prevalence with lactulose and trehalose ingestion, and the effect of lactulose was significantly stronger than that of trehalose at the same dose level (p < 0.05). The quantity of trehalose and lactulose that induced diarrhea differed greatly from person to person. The transitory laxative threshold was estimated as 0.65 g/kg body weight for trehalose and 0.26 g/kg body weight for lactulose by using the regression equation between the dose levels of the test substances and the cumulative incidence of diarrhea. These results suggest that it would be quite acceptable to administer trehalose up to 33 g and lactulose up to 13 g in a person weighing 50 kg.

Abdomen↗
PubMed2000

[Studies on the trehalose-producing by Colloides CB39].

A bacterium which had the ability of producing sugar produced sugar at low temperature (18 degrees C) was selected from the water of Tianchi in Changbai mountain. The sugar was identified to be trehalose by methods of thin-layer chromatography, reaction of producing osazyone, capillary electrophoresis and infrared spectrum. It was also been found that the trehalose produced by this bacterium (identified to be a new species of Colloides Sp. CB39) was exocrine. At 18 degrees C, its trehalose content in culture solution was 256 mg/g dry weight of the strain. This characteristic is different from that of other strains, which had been proved to have the ability of producing trehalose. then the strain CB39 was induced by ultraviolet in order to abstain mutants which can produce trehalose in high level at usual temperature. The mutant strain 5 that could produce trehalose in high level at usual temperature (25 degrees C) was selected from all the mutant strains. The value of its productivity of trehalose is 416.7 mg/g dry weight of the strain. (At the same temperature, trehalose productivity of mutant strain 5 was eight times of that of the wild type strain CB39).

Bacteria↗
PubMed2002

Substrate-binding recognition and specificity of trehalose phosphorylase from Schizophyllum commune examined in steady-state kinetic studies with deoxy and deoxyfluoro substrate analogues and inhibitors.

Trehalose phosphorylase is a component of the alpha-D-glucopyranosyl alpha-D-glucopyranoside (alpha,alpha-trehalose)-degrading enzyme system in fungi and it catalyses glucosyl transfer from alpha,alpha-trehalose to phosphate with net retention of the anomeric configuration. The enzyme active site has no detectable affinity for alpha,alpha-trehalose in the absence of bound phosphate and catalysis occurs from the ternary complex. To examine the role of non-covalent enzyme-substrate interactions for trehalose phosphorylase recognition, we used the purified enzyme from Schizophyllum commune and tested a series of incompetent structural analogues of the natural substrates and products as inhibitors of the enzyme. Equilibrium-binding constants (K(i)) for deoxy- and deoxyfluoro derivatives of D-glucose show that loss of interactions with the 3-, 4- or 6-OH, but not the reactive 1- and the 2-OH, results in considerably (> or =100-fold) weaker affinity for sugar-binding subsite +1, revealing the requirement for hydrogen bonding with hydroxyls, away from the site of chemical transformation to position precisely the D-glucose-leaving group/nucleophile for catalysis. The high specificity of trehalose phosphorylase for the sugar aglycon during binding and conversion of O-glycosides is in contrast with the observed alpha-retaining phosphorolysis of alpha-D-glucose-1-fluoride (alpha-D-Glc-1-F) since the productive bonding capability of the fluoride-leaving group with subsite +1 is minimal. The specificity constant (19 M(-1).s(-1)) and catalytic-centre activity (0.1 s(-1)) for the reaction with alpha-D-Glc-1-F are 0.10- and 0.008-fold the corresponding kinetic parameters for the enzymic reaction with alpha,alpha-trehalose. The non-selective-inhibition profile for a series of inactive alpha-D-glycopyranosyl phosphates shows that the driving force for the binary-complex formation lies mainly in interactions of the enzyme with the phosphate group and suggests that hydrogen bonding with hydroxyl groups at the catalytic site (subsite -1) contributes to catalysis by providing stabilization, which is specific to the transition state. Vanadate, a tight-binding phosphate mimic, inhibits the phosphorolysis of alpha-D-Glc-1-F by forming a ternary complex whose apparent dissociation constant of 120 microM is approx. 160-fold greater than the dissociation constant of the same inhibitor complex with alpha,alpha-trehalose.

Arsenates↗
PubMed2002

Purification, cloning, expression, and properties of mycobacterial trehalose-phosphate phosphatase.

The trehalose-phosphate phosphatase (TPP) was purified from the cytosol of Mycobacterium smegmatis to near homogeneity using a variety of conventional steps to achieve a purification of about 1600-fold with a yield of active enzyme of about 1%. Based on gel filtration, the active enzyme had a molecular weight of about 27,000, and the most purified fraction also gave a major band on SDS-PAGE corresponding to a molecular weight of about 27,000. A number of peptides from the 27-kDa protein were sequenced and these sequences showed considerable homology to the trehalose-P phosphatase (otsB) of Escherichia coli. Based on these peptides, the M. smegmatis gene for TPP was cloned and expressed in E. coli. The recombinant protein was synthesized with a (His)(6) tag at the amino terminus. Most of the TPP activity in the crude E. coli sonicate was initially found in the membrane fraction, but it became solubilized in the presence of 0.2% Sarkosyl. The solubilized protein was purified to apparent homogeneity on a metal ion column and this fraction had high phosphatase activity that was completely specific for trehalose-P. The purified enzyme, either isolated from M. smegmatis, or expressed in E. coli, rapidly dephosphorylated trehalose-6-P, but had essentially no activity on any other sugar phosphates, or on p-nitrophenyl phosphate. The K(m) for trehalose-6-P was about 1.6 mm, and the pH optimum was about 7.5. The native enzyme showed an almost absolute requirement for Mg(2+) and was not very active with Mn(2+), whereas both of these cations were equally effective with the recombinant TPP. The enzyme activity was inhibited by the antibiotics, diumycin and moenomycin, but not by a number of other antibiotics or trehalose analogs. TPP activity was strongly inhibited by the detergents, Sarkosyl and deoxycholate, even at 0.025%, but it was not inhibited by Nonidet P-40, Triton X-100, or octyl glucoside, even at concentrations up to 0.3%. The purified enzyme was stable to heating at 60 degrees C for up to 6 min, but was slowly inactivated at 70 degrees C. Circular dichroism studies on recombinant TPP indicate that the secondary structure of this protein has considerable beta-pleated sheet and is very compact. TPP may play a key role in the biosynthesis of trehalose compounds, such as trehalose mycolates, and therefore may represent an excellent target site for chemotherapy against tuberculosis and other mycobacterial diseases.

Amino Acid Sequence↗
PubMed1986

Enzymes of alpha,alpha-Trehalose Metabolism in Soybean Nodules.

Metabolism of trehalose, alpha,d-glucopyranosyl-alpha,d-glucopyranoside, was studied in nodules of Bradyrhizobium japonicum-Glycine max [L.] Merr. cv Beeson 80 symbiosis. The nodule extract was divided into three fractions: bacteroid soluble protein, bacteroid fragments, and cytosol. The bacteroid soluble protein and cytosol fractions were gel-filtered. The key biosynthetic enzyme, trehalose-6-phosphate synthetase, was consistently found only in the bacteroids. Trehalose-6-phosphate phosphatase activity was present both in the bacteroid soluble protein and cytosol fractions. Trehalase, the most abundant catabolic enzyme was present in all three fractions and showed two pH optima: pH 3.8 and 6.6. Two other degradative enzymes, phosphotrehalase, acting on trehalose-6-phosphate forming glucose and glucose-6-phosphate, and trehalose phosphorylase, forming glucose and beta-glucose-1-phosphate, were also detected in the bacteroid soluble protein and cytosol fractions. Trehalase was present in large excess over trehalose-6-phosphate synthetase. Trehalose accumulation in the nodules would appear to be predicated on spatial separation of trehalose and trehalase.

Journal Article↗