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PubMed1994

The influence of altered gravity on carbohydrate metabolism in excised wheat leaves.

We developed a system to study the influence of altered gravity on carbohydrate metabolism in excised wheat leaves by means of clinorotation. The use of excised leaves in our clinostat studies offered a number of advantages over the use of whole plants, most important of which were minimization of exogenous mechanical stress and a greater amount of carbohydrate accumulation during the time of treatment. We found that horizontal clinorotation of excised wheat leaves resulted in significant reductions in the accumulation of fructose, sucrose, starch and fructan relative to control, vertically clinorotated leaves. Photosynthesis, dark respiration and the extractable activities of ADP glucose pyrophosphorylase (EC 2.7.7.27), sucrose phosphate synthase (EC 2.4.4.14), sucrose sucrose fructosyltransferase (EC 2.4.1.99), and fructan hydrolase (EC 3.2.1.80) were unchanged due to altered gravity treatment.

Bacterial Proteins↗

PubMed2004

Streptococcus mutans fructosyltransferase interactions with glucans.

Streptococcus mutans utilizes sucrose to synthesize glucans by glucosyltransferase and fructans by fructosyltransferase (FTF). Antibodies raised against a recombinant FTF were used to study S. mutans FTF secretion. Low amounts of cell-free FTF were found in culture of S. mutans grown with sucrose, while an increase in bacteria displaying cell surface FTF was detected. FTF added to S. mutans cultures was adsorbed to bacteria grown with sucrose but not to bacteria grown with glucose or fructose or to a gtf inactivated mutant grown with sucrose. Recombinant FTF was found to have high affinity for glucans suggesting that fructans and glucans are an integral part of the polysaccharide matrix of oral biofilms.

Biofilms↗

PubMed2002

In vitro bulb development in shallot (Allium cepa L. Aggregatum Group): effects of anti-gibberellins, sucrose and light.

Bulbing was studied in shallot plants cultured in vitro. Bulbing occurred under a 16 h photoperiod with fluorescent + incandescent light and 30-50 g 1(-1) sucrose in the culture medium. Exogenous gibberellin (10 microM GA3) inhibited leaf and root growth and bulbing. When added to the medium at a concentration of 10 microM, three inhibitors of gibberellin biosynthesis (ancymidol, flurprimidol and paclobutrazol) promoted bulb formation and the percentage of bulbing. When ancymidol was used in combination with GA3, it did not reverse the effect of GA3 applied alone. Under treatments with 30-70 g l(-1) sucrose, bulbing ratios greater than those found in control plants were achieved by addition of ancymidol, and bulb fresh weight was increased in the same way. Ancymidol caused a 66% decrease in sucrose content in leaf bases but greatly increased the glucose, fructose and fructan contents. The increase in fructan content by ancymidol could result from the three-fold rise in total [14C]sucrose uptake per plant from the culture medium associated with a marked increase in leaf base labelling at the expense of root labelling. The possible role of ancymidol is discussed and evidence supports a major regulatory role for gibberellins in bulbing.

Allium↗

PubMed2006

The levansucrase and inulosucrase enzymes of Lactobacillus reuteri 121 catalyse processive and non-processive transglycosylation reactions.

Bacterial fructosyltransferase (FTF) enzymes synthesize fructan polymers from sucrose. FTFs catalyse two different reactions, depending on the nature of the acceptor, resulting in: (i) transglycosylation, when the growing fructan chain (polymerization), or mono- and oligosaccharides (oligosaccharide synthesis), are used as the acceptor substrate; (ii) hydrolysis, when water is used as the acceptor. Lactobacillus reuteri 121 levansucrase (Lev) and inulosucrase (Inu) enzymes are closely related at the amino acid sequence level (86 % similarity). Also, the eight amino acid residues known to be involved in catalysis and/or sucrose binding are completely conserved. Nevertheless, these enzymes differ markedly in their reaction and product specificities, i.e. in beta(2-->6)- versus beta(2-->1)-glycosidic-bond specificity (resulting in levan and inulin synthesis, respectively), and in the ratio of hydrolysis versus transglycosylation activities [resulting in glucose and fructooligosaccharides (FOSs)/polymer synthesis, respectively]. The authors report a detailed characterization of the transglycosylation reaction products synthesized by the Lb. reuteri 121 Lev and Inu enzymes from sucrose and related oligosaccharide substrates. Lev mainly converted sucrose into a large levan polymer (processive reaction), whereas Inu synthesized mainly a broad range of FOSs of the inulin type (non-processive reaction). Interestingly, the two FTF enzymes were also able to utilize various inulin-type FOSs (1-kestose, 1,1-nystose and 1,1,1-kestopentaose) as substrates, catalysing a disproportionation reaction; to the best of our knowledge, this has not been reported for bacterial FTF enzymes. Based on these data, a model is proposed for the organization of the sugar-binding subsites in the two Lb. reuteri 121 FTF enzymes. This model also explains the catalytic mechanism of the enzymes, and differences in their product specificities.

Binding Sites↗

PubMed1993

Sucrose-derived exopolysaccharides of Streptococcus mutans V403 contribute to infectivity in endocarditis.

We used an isogenic mutant of Streptococcus mutans V403, which differs from the wild-type V403 in genes involved in glucan and fructan production, to examine the importance of these exopolysaccharides as factors affecting infectivity in endocarditis. Rats inoculated with V403 developed endocarditis more frequently than animals inoculated with the mutant strain which produced neither glucan nor fructan (58% versus 12%, P < 0.01). In phagocytosis assays, both strains were found to be associated with the human granulocytes but a greater number of live V403 than of mutant organisms could be recovered. Colony counts recovered from fibrin plates incubated with the mutant were lower than those incubated with V403. These experiments indicate that exopolysaccharides produced by Streptococcus mutans contribute to its infectivity in endocarditis.

Alleles↗

PubMedDate not supplied

Importance of enzyme purity and activity in the measurement of total dietary fiber and dietary fiber components.

A study was made of the effect of the activity and purity of enzymes in the assay of total dietary fiber (AOAC Method 985.29) and specific dietary fiber components: resistant starch, fructan, and beta-glucan. In the measurement of total dietary fiber content of resistant starch samples, the concentration of alpha-amylase is critical; however, variations in the level of amyloglucosidase have little effect. Contamination of amyloglucosidase preparations with cellulase can result in significant underestimation of dietary fiber values for samples containing beta-glucan. Pure beta-glucan and cellulase purified from Aspergillus niger amyloglucosidase preparations were used to determine acceptable critical levels of contamination. Sucrose, which interferes with the measurement of inulin and fructooligosaccharides in plant materials and food products, must be removed by hydrolysis of the sucrose to glucose and fructose with a specific enzyme (sucrase) followed by borohydride reduction of the free sugars. Unlike invertase, sucrase has no action on low degree of polymerization (DP) fructooligosaccharides, such as kestose or kestotetraose. Fructan is hydrolyzed to fructose and glucose by the combined action of highly purified exo- and endo-inulinases, and these sugars are measured by the p-hydroxybenzoic acid hydrazide reducing sugar method. Specific measurement of beta-glucan in cereal flour and food extracts requires the use of highly purified endo-1,3:1,4 beta-glucanase and A. niger beta-glucosidase. Beta-glucosidase from almonds does not completely hydrolyze mixed linkage beta-glucooligosaccharides from barley or oat beta-glucan. Contamination of these enzymes with starch, maltosaccharide, or sucrose-hydrolyzing enzymes results in production of free glucose from a source other than beta-glucan, and thus an overestimation of beta-glucan content. The glucose oxidase and peroxidase used in the glucose determination reagent must be essentially devoid of catalase and alpha- and beta-glucosidase.

Aspergillus niger↗

PubMedDate not supplied

Measurement of novel dietary fibers.

With the recognition that resistant starch (RS) and nondigestible oligosaccharides (NDO) act physiologically as dietary fiber (DF), a need has developed for specific and reliable assay procedures for these components. The ability of AOAC DF methods to accurately measure RS is dependent on the nature of the RS being analyzed. In general, NDO are not measured at all by AOAC DF Methods 985.29 or 991.43, the one exception being the high molecular weight fraction of fructo-oligosaccharides. Values obtained for RS, in general, are not in good agreement with values obtained by in vitro procedures that more closely imitate the in vivo situation in the human digestive tract. Consequently, specific methods for the accurate measurement of RS and NDO have been developed and validated through interlaboratory studies. In this paper, modifications to AOAC fructan Method 999.03 to allow accurate measurement of enzymically produced fructo-oligosaccharides are described. Suggested modifications to AOAC DF methods to ensure complete removal of fructan and RS, and to simplify pH adjustment before amyloglucosidase addition, are also described.

Dietary Fiber↗

PubMed2003

Catalytic activity of zymomonas mobilis extracellular "levan-levansucrase" complex in sucrose medium.

The fructan biosynthesis by ethanol sedimented "levan-levansucrase" complex from Zymomonas mobilis fermentation broth as well as purified levansucrase was investigated. The fructooligosaccharide (FOS) producing activity of "levan-levamsucrase" sediment was investigated in 55% sucrose syrup at 45 degrees C. It was shown that FOS in the syrup were presented by 1-kestose, 6-kestose, neokestose and nystose. The increase of gluconic acid concentration was observed in the reaction mixture during the incubation suggesting about presence of glucose/fructose oxidoreductase in "levan-levansucrase" sediment. The influence of ethanol, glycerol and NaCl on levan and fructooligosaccharide formation by "levan-levansucrase" complex and purified levansucrase was studied and the changes in the ratio between different activities of levansucrase (sucrose hydrolysis, levan biosynthesis and FOS formation) were observed. Ethanol increases the FOS biosynthesis part in total activity of purified levansucrase. The technology of the production of prebiotics containing food product--fructan syrup by "levan-levansucrase " sediment as biocatalyst was developed.

Catalysis↗

PubMed1990

Isolation and some properties of extracellular glucan-producing strains of human oral Streptococcus salivarius.

A total of eighteen strains of Streptococcus salivarius, which formed rough gelatinous, rough mucoid or smooth mucoid colonies on sucrose agar media, were isolated from the saliva and tongue dorsum of adults. All of the isolates produced glucans as well as fructans from sucrose. The bulk of the glucans was synthesized by the extracellular enzyme fraction and was water insoluble, whereas most of the fructans were synthesized by the cell-associated enzyme fraction and were water soluble. All strains formed microbial deposits on wire and glass surfaces when cultured in sucrose broth, but their sucrose-dependent adhesion was apparently looser than that produced by a cariogenic S. sobrinus strain. The rough gelatinous colony forming strains possessed a greater ability to synthesize water-insoluble glucans and produced heavier deposits with higher cohesion. Preliminary studies showed that the S. salivarius of such characteristic forms of colony were detected primarily in the saliva and tongue dorsum: the smooth mucoid colony formers appeared to predominate in the tongue coat and the rough mucoid and rough gelatinous colony formers were prominent in saliva. Isolation of these S. salivarius from dental plaques was low.

Adult↗

PubMed1987

[The structure of O-specific polysaccharide isolated from the lipopolysaccharide of Yersinia intermedia strain 180].

An O-specific polysaccharide from lipopolysaccharide of Yersinia intermedia pathogenic strain 680 has been isolated and shown to be a serologically active fructane. Serological specificity of the lipopolysaccharide and the fructane was studied by reactions of precipitation and of inhibition of passive hemolysis. On the basis of methylation studies, 13C NMR spectroscopy, and immunochemical data the following structure was proposed for the repeating unit of the O-specific polysaccharide: (Formula: see text)

Antigens, Bacterial↗

PubMed2002

Applications of inulin and oligofructose in health and nutrition.

Inulin and oligofructose belong to a class of carbohydrates known as fructans. The main sources of inulin and oligofructose that are used in the food industry are chicory and Jerusalem artichoke. Inulin and oligofructose are considered as functional food ingredients since they affect the physiological and biochemical processes in rats and human beings, resulting in better health and reduction in the risk of many diseases. Experimental studies have shown their use as bifidogenic agents, stimulating the immune system of the body, decreasing the pathogenic bacteria in the intestine, relieving constipation, decreasing the risk of osteoporosis by increasing mineral absorption, especially of calcium, reducing the risk of atherosclerosis by lowering the synthesis of triglycerides and fatty acids in the liver and decreasing their level in serum. These fructans modulate the hormonal level of insulin and glucagon, thereby regulating carbohydrate and lipid metabolism by lowering the blood glucose levels; they are also effective in lowering the blood urea and uric acid levels, thereby maintaining the nitrogen balance. Inulin and oligofructose also reduce the incidence of colon cancer. The biochemical basis of these beneficial effects of inulin and oligofructose have been discussed. Oligofructose are non cariogenic as they are not used by Streptococcus mutans to form acids and insoluble glucans that are the main culprits in dental caries. Because of the large number of health promoting functions of inulin and oligofructose, these have wide applications in various types of foods like confectionery, fruit preparations, milk desserts, yogurt and fresh cheese, baked goods, chocolate, ice cream and sauces. Inulin can also be used for the preparation of fructose syrups.

Animals↗

PubMed2004

Absence of turnover and futile cycling of sucrose in leaves of Lolium temulentum L.: implications for metabolic compartmentation.

To study the interdependence of sucrose accumulation and its hydrolyzing enzyme, soluble acid invertase (AI; EC 3.2.1.26), in fructan-accumulating temperate grasses and cereals, experiments were performed in which sucrose synthesis was abolished in leaves of Lolium temulentum by four independent inhibitory factors, each having a distinct mechanism of action. Trials in the light with mannose or vanadate and in the dark with anoxia or cyanide showed that previously accumulated sucrose was stable in the tissue over a 5- to 6-h period. Conversely, putatively vacuolar AI activity in tissue homogenates was sufficient to completely convert endogenous sucrose to monosaccharide within the same period. Continuous invertase-mediated breakdown of sucrose was thus not a feature of this tissue. It is concluded that AI and sucrose were not in metabolic contact in vivo, implying differential compartmentation. In darkness, in uninhibited leaves, sucrose concentrations fell linearly with respect to time at a rate of -0.6 mg g(-1) FW h(-1), over a 5- to 6-h period. This value is equivalent to rates of dark respiration measured by gas exchange. Dark-utilisation of sucrose was not accompanied by monosaccharide accumulation in the tissue. The rate of sucrose loss was 3-fold lower than rates of extractable AI activity. Hence, if AI was involved in dark-utilisation, then this implies at least a partial differential localisation of enzyme and substrate. However, the dark-consumption of sucrose was completely abolished by anoxia and by cyanide. It follows that dark-mobilisation (unlike invertase hydrolysis per se) was respiration-dependent and did not result from a simple co-localisation of sucrose and invertase. Taken together, the results show that sucrose and invertase do not share the same metabolic compartment in grass leaves. It is possible that invertase has no role in the mobilisation of stored sucrose in leaves of the fructan-accumulating grasses.

Darkness↗

PubMed2004

Performance of a generalist grasshopper on a C3 and a C4 grass: compensation for the effects of elevated CO2 on plant nutritional quality.

The increasing CO2 concentration in Earth's atmosphere is expected to cause a greater decline in the nutritional quality of C3 than C4 plants. As a compensatory response, herbivorous insects may increase their feeding disproportionately on C3 plants. These hypotheses were tested by growing the grasses Lolium multiflorum C3) and Bouteloua curtipendula C4) at ambient (370 ppm) and elevated (740 ppm) CO2 levels in open top chambers in the field, and comparing the growth and digestive efficiencies of the generalist grasshopper Melanoplus sanguinipes on each of the four plant x CO2 treatment combinations. As expected, the nutritional quality of the C3 grass declined to a greater extent than did that of the C4 grass at elevated CO2; protein levels declined in the C3 grass, while levels of carbohydrates (sugar, fructan and starch) increased. However, M. sanguinipes did not significantly increase its consumption rate to compensate for the lower nutritional quality of the C3 grass grown under elevated CO2. Instead, these grasshoppers appear to use post-ingestive mechanisms to maintain their growth rates on the C3 grass under elevated CO2. Consumption rates of the C3 and C4 grasses were also similar, demonstrating a lack of compensatory feeding on the C4 grass. We also examined the relative efficiencies of nutrient utilization from a C3 and C4 grass by M. sanguinipes to test the basis for the C4 plant avoidance hypothesis. Contrary to this hypothesis, neither protein nor sugar was digested with a lower efficiency from the C4 grass than from the C3 grass. A novel finding of this study is that fructan, a potentially large carbohydrate source in C3 grasses, is utilized by grasshoppers. Based on the higher nutrient levels in the C3 grass and the better growth performance of M. sanguinipes on this grass at both CO2 levels, we conclude that C3 grasses are likely to remain better host plants than C4 grasses in future CO2 conditions.

Adaptation, Physiological↗

PubMed2005

Effect of the carbohydrate composition of feed concentratates on methane emission from dairy cows and their slurry.

Dietary carbohydrate effects on methane emission from cows and their slurry were measured on an individual animal basis. Twelve dairy cows were fed three of six diets each (n = 6 per diet) of a forage-to-concentrate ratio of 1 : 1 (dry matter basis), and designed to cover the cows' requirements. The forages consisted of maize and grass silage, and hay. Variations were exclusively accomplished in the concentrates which were either rich in lignified or non-lignified fiber, pectin, fructan, sugar or starch. To measure methane emission, cows were placed into open-circuit respiration chambers and slurry was stored for 14 weeks in 60-L barrels with slurry being intermittently connected to this system. The enteric and slurry organic matter digestibility and degradation was highest when offering Jerusalem artichoke tubers rich in fructan, while acid-detergent fiber digestibility and degradation were highest in cows and slurries with the soybean hulls diet rich in non-lignified fiber. Multiple regression analysis, based on nutrients either offered or digested, suggested that, when carbohydrate variation is done in concentrate, sugar enhances enteric methanogenesis. The methane emission from the slurry accounted for 16.0 to 21.9% of total system methane emission. Despite a high individual variation, the methane emission from the slurry showed a trend toward lower values, when the diet was characterized by lignified fiber, a diet where enteric methane release also had been lowest. The study disproved the assumption that a lower enteric methanogenesis, associated with a higher excretion of fiber, will inevitably lead to compensatory increases in methane emission during slurry storage.

Animal Feed↗

PubMed1996

Inulin synthesis by a combination of purified fructosyltransferases from tubers of Helianthus tuberosus.

Sucrose-sucrose 1-fructosyltransferase (1-SST) was purified 100-fold from tubers of Helianthus tuberosus L. The purified enzyme was essentially devoid of invertase activity and could be separated by isoelectric focusing into five isoforms which all were composed of two subunits (59 and 26 kDa). Fructan-fructan 1-fructosyltransferase (1-FFT) was purified from the same source [M. Lüscher et al. (1993) New Phytologist 123, 437-442). When incubated individually with sucrose, 1-FFT was inactive while 1-SST formed isokestose (trimer) and, upon prolonged incubation, some nystose (tetramer). When a combination of the two enzymes was incubated with sucrose, a series of oligofructosides with a degree of polymerization of up to 20 was formed. Amino acid sequences of tryptic peptide fragments from both 1-SST and 1-FFT indicate that these enzymes are highly homologous with plant invertases.

Amino Acid Sequence↗

PubMed2006

Monosaccharide composition, chain length and linkage type influence the interactions of oligosaccharides with dry phosphatidylcholine membranes.

Sugars play an important role in the desiccation tolerance of most anhydrobiotic organisms and disaccharides have been extensively investigated for their ability to stabilize model membranes in the dry state. Much less is known about the ability of oligosaccharides to protect dry membranes. However, it has been shown that different structural families of oligosaccharides have different efficacies to interact with and protect membranes during drying. Here, we have compared three families of linear oligosaccharides (fructans, malto-oligosaccharides, manno-oligosaccharides) for their chain-length dependent lyoprotective effect on egg phosphatidylcholine liposomes. We found increased protection with chain length for the fructans, a moderate decrease in protection with chain length for malto-oligosaccharides, and a strong decrease for manno-oligosaccharides. Using Fourier-transform infrared spectroscopy and differential scanning calorimetry, we show that the degree of lyoprotection of the different sugars is closely related to their influence on the gel to liquid-crystalline phase behavior of the dry membranes and to the extent of H-bonding to different groups (C=O, P=O, choline) in the lipids. Possible structural characteristics of the different oligosaccharides that may determine the extent to which they are able to interact with and protect membranes are discussed.

Calorimetry, Differential Scanning↗

PubMed2006

Effect of oxazaborolidines on immobilized fructosyltransferase analyzed by surface plasmon resonance.

Dental diseases are among the most prevalent afflictions of humankind. These diseases are associated with the formation of biofilms harboring pathogenic bacteria. Fructosyltransferases (FTF) are extra cellular enzymes of several oral bacteria. FTF are associated with the formation of extracellular polysaccharide matrix (fructans) which play a role in biofilm formation and oral bacteria physiology. Oxazaborolidines have been shown to inhibit biofilm formation. The purpose of this study was to examine if the anti-biofilm effect is, in part, an effect on the immobilized enzymes synthesizing the extra cellular polysaccharide participating in biofilm formation. Eight different oxazaborolidines (BNO1-BNO8) were synthesized and evaluated for their affect on the synthesis of fructans by FTF using the biomolecular interaction analysis (BIAcore) system which involves the use of real-time surface plasmon resonance (SPR) technique. The tested oxazaborolidines demonstrated a significant and immediate inhibitory effect on immobilized FTF activity. This effect was reversible. Our results show that oxazaborolidines can act as enzymatic inhibitors of FTF immobilized on the surface, also at levels lower than their MIC. Part of the anti-biofilm effect of BNOs may be accounted for this enzymatic inhibition.

Anti-Bacterial Agents↗

PubMed2004

Effect of carbohydrates on fructosyltransferase expression and distribution in Streptococcus mutans GS-5 biofilms.

Streptococcus mutans produces a fructosyltransferase (FTF) enzyme, which synthesizes fructan polymers from sucrose. Fructans contribute to the virulence of the biofilm by acting as binding sites for S. mutans adhesion and as extracellular nutrition reservoir for the oral bacteria. Antibodies raised against a recombinant S. mutans FTF were used to test the effect of glucose, fructose, and sucrose on FTF expression in S. mutans GS-5 biofilms. Biofilms formed in the presence of fructose and glucose showed a higher ratio of FTF compared to biofilms formed in the presence of sucrose. Confocal laser scanning microscopy images of S. mutans biofilms indicated a carbohydrate-dependent FTF distribution. The layer adjacent to the surface and those at the liquid interface displayed high amounts cell-free FTF with limited amount of bacteria while the in-between layers demonstrated both cell-free FTF and cells expressing cell-surface FTF. Biofilm of S. mutans grown on hydroxyapatite surfaces expressed several FTF bands with molecular masses of 160, 125, 120, 100, and 50 kDa, as detected by using FTF specific antibodies. The results show that FTF expression and distribution in S. mutans GS-5 biofilms is carbohydrate regulated.

Antibodies, Bacterial↗