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Prebiotics: preferential substrates for specific germs?

A prebiotic is "a non-digestible food ingredient that beneficially affects the host by selectively stimulating the growth and/or the activity of one or a limited number of bacteria in the colon." The premise is based on the hypothesis that the large gut in humans contains bacteria that are beneficial or detrimental to health. Although this generalization probably gives too simplistic a view of gut microbiology, it is a feasible working concept. Currently, food components that seem to exert the best prebiotic effects are inulin-type fructans. In pure culture, most species of bifidobacteria are adapted to the utilization of these nondigestible oligosaccharides but many other bacteria are also capable of metabolizing them. Clearly, these studies of pure bacteria are of limited use unless their results are supported by the results of studies using mixed cultures. Indeed, as many components of the gut microbiota as possible should be measured to indicate a true prebiotic effect. Simple stimulation of bifidobacteria is insufficient to demonstrate an effect; the effects on other gut microorganisms in vivo with human volunteers is necessary. Adjustment of the composition and activities of the colonic microflora so that health-promoting activities are optimized remains key in functional food development. New methods are being applied extensively to human gut microbiology and promise the degree of reliability required to detect subtle changes in colonic microflora composition and to correlate such changes with health benefits. This is a review of the present state of knowledge concerning prebiotics, with emphasis on the criteria used for classification, mechanisms of selective growth stimulation, and physiologic effects.

Bifidobacterium↗

Carbohydrate concentrations in crown fractions from winter oat during hardening at sub-zero temperatures.

BACKGROUND AND AIMS: Contradictory results in correlation studies of plant carbohydrates with freezing tolerance may be because whole crown tissue is analysed for carbohydrates while differences exist in the survival of specific tissue within the crown. The aim of this study was to see if carbohydrate changes in tissue within oat crowns during second phase hardening (sub-zero hardening) are tissue specific. METHODS: The lower portion of oat (Avena sativa) crowns was exposed to mild grinding in a blender and the remaining crown meristem complex, consisting of tough root-like vessels, was ground in a device developed specifically for grinding cereal crown tissue. Carbohydrates were extracted by water and measured by HPLC. Carbohydrate concentrations were compared in the two regions of the crown before and after hardening at sub-zero temperatures. KEY RESULTS: Fructan of all size classes except DP>6 decreased during sub-zero hardening in both stems (base of leaf sheath) and crown meristem complex. Total simple sugar increase, including sucrose, was significantly higher in the crown meristem complex than in the stem. CONCLUSIONS: Results support the hypothesis that carbohydrate change in mildly frozen plants is tissue specific within crowns and underscore the need to evaluate specific tissue within the crown when correlating the biochemistry of plants with freezing tolerance.

Adaptation, Physiological↗

Beef induces and rye bran prevents the formation of intestinal polyps in Apc(Min) mice: relation to beta-catenin and PKC isozymes.

Epidemiological studies suggest that high consumption of red meat and saturated fat and low consumption of fiber are associated with an increased risk of colon cancer. Therefore, we studied whether diets high in red meat or high in different grain fibers as well as inulin, polydisperse beta(2-->1) fructan, could affect the formation of intestinal polyps in Apc(Min) mice. Min mice were fed the following high-fat (40% of energy) diets for 5-6 weeks; a high-beef diet and a casein-based diet without added fiber or casein-based diet with 10% (w/w) oat, rye or wheat bran, or 2.5% (w/w) inulin. One group had a normal low-fat AIN93-G diet. The mice fed the rye-bran diet had the lowest number of polyps in the distal small intestine [15.4 +/- 8.7 (mean +/- SD)], and in the entire intestine (26.4 +/- 12.1). The rye-bran group differed significantly (P = 0. 001-0.004) from the beef group (36.6 +/- 9.4 and 52.8 +/- 13.2). In addition, the beef group differed significantly from the AIN93-G group (P = 0.009) and also from the wheat-bran group (21.0 +/- 6.1 and 35.0 +/- 8.2; P = 0.02) in the distal small intestine. The inulin group (32.9 +/- 14.3 and 49.3 +/- 16.3), on the other hand, was close to the beef group and it differed significantly from the rye-bran group in the distal small intestine. The number of animals bearing tumors in the colon + caecum was only 33% in the rye-bran group when compared with 89% in the beef and 100% in the inulin groups. The mice fed the rye-bran and beef diets had the lowest levels of cytosolic beta-catenin (0.60 +/- 0.42 and 0.67 +/- 0.26) and they differed significantly (P = 0.040 and 0.062) from the mice fed the oat-bran diet (1.46 +/- 0.43). No differences between groups in expression of protein kinase C (PKC) alpha, betaII, delta and zeta were found. The four PKC isozymes were positively correlated with cytosolic beta-catenin levels (r = 0.62-0.68; P < 0.0001).

Animals↗

Antitumorigenic activity of the prebiotic inulin enriched with oligofructose in combination with the probiotics Lactobacillus rhamnosus and Bifidobacterium lactis on azoxymethane-induced colon carcinogenesis in rats.

Prebiotics such as fructans, and probiotics such as Lactobacilli or Bifidobacteria, or a combination of prebiotics and probiotics (synbiotics) are thought to be protective against colon cancer. Therefore, we studied whether the prebiotic inulin enriched with oligofructose (Raftilose-Synergy1, briefly, Synergy1, 10% of the diet), probiotics [Bifidobacterium lactis (Bb12) and Lactobacillus rhamnosus (LGG), each at 5x10(8) c.f.u./g diet] or synbiotics (a combination of the two) protect rats against azoxymethane (AOM)-induced colon cancer. Male F344 rats were divided into: Controls; PRE, which were fed a diet containing Synergy1; PRO, fed a diet containing LGG and Bb12; PREPRO, fed a diet containing Synergy1, LGG and BB12. Ten days after beginning the diets, rats were treated with AOM (15 mg/kg s.c. two times); dietary treatments were continued for the entire experiment. Thirty-one weeks after AOM, rats treated with Synergy1 (PRE and PREPRO groups) had a significantly lower (P < 0.001) number of tumours (adenomas and cancers) than rats without Synergy1 (colorectal tumours/rat were 1.9 +/- 1.7, 1.1 +/- 1.1, 2.2 +/- 1.4 and 0.9 +/- 1.2 in Controls, PRE, PRO and PREPRO groups, respectively, means +/- SD). A slight, not significant effect of probiotics in reducing malignant tumours was also observed (P = 0.079). Caecal short-chain fatty acids (SCFA) were higher (P < 0.001) in the groups treated with Synergy1. Apoptosis was increased in the normal mucosa of the PRO group, while no variation was observed in the tumours. Colonic proliferation was lower in the PRE group as compared with Controls. Glutathione S-transferase placental enzyme pi type expression, and to a lesser extent, inducible NO synthase were depressed in the tumours from rats in the PRE and PREPRO groups. Cycloxygenase-2 expression was increased in the tumours of control rats but not in those from PRE, PRO or PREPRO rats. In conclusion, prebiotic administration in the diet decreases AOM-induced carcinogenesis in rats.

Adenocarcinoma↗

Inhibition of Streptococcus mutans biofilm accumulation and polysaccharide production by apigenin and tt-farnesol.

OBJECTIVES: Apigenin is a potent inhibitor of glucosyltransferases and tt-farnesol affects the membrane integrity of Streptococcus mutans. We investigated the influence of apigenin and tt-farnesol, alone and in combination, on the accumulation, polysaccharide composition and viability of S. mutans UA159 biofilms. METHODS: Initially, biofilms were grown for 54 h; then, the early-formed biofilms were treated for 1 min twice daily with one of the following: (i). 1.33 mM tt-farnesol; (ii). 1.33 mM apigenin; (iii). apigenin + tt-farnesol (1.33 mM each); (iv). vehicle control (20% ethanol with 0.75% dimethyl sulphoxide); (v). 0.12% chlorhexidine (1.33 mM); or (vi). physiological saline (145 mM NaCl). The procedure was repeated at biofilm ages of 78 and 102 h, and biofilms were harvested at 126 h. The dry weight, protein concentration, number of cfu, and polysaccharide composition per biofilm were determined. RESULTS: The dry weights of the biofilms treated with the test agents were significantly less (30-50%) than those treated with vehicle control (P < 0.05). Biofilms treated with the test agents also resulted in lower amounts of extracellular alkali-soluble glucans, intracellular iodophilic polysaccharides and, to a lesser extent, fructans. The fructosyltransferase activity was affected only by apigenin and apigenin + tt-farnesol. The recoverable viable counts of S. mutans were slightly lower (0.5 to 1 log10 decrease in cfu/biofilm) after apigenin and tt-farnesol treatments compared with the vehicle control. Chlorhexidine displayed potent bactericidal activity, and virtually halted the further accumulation of early-formed (54 h old) biofilms. CONCLUSIONS: Apigenin and tt-farnesol affected the accumulation and polysaccharide content of S. mutans biofilms without major impact on the bacterial viability.

Anti-Infective Agents, Local↗

Concepts in functional foods: the case of inulin and oligofructose.

Recent advances in biosciences support the hypothesis that diet modulates various body functions. Diet may maintain well-being and reduce the risk of some diseases. Such discoveries have led to the concept of "functional food" and the development of the new discipline, i.e., "functional food science." A practical and simple definition of a "functional food" is a food for which a claim has been authorized. The food components to be discussed as potential "functional food ingredients" are the inulin-type fructans, i.e., chicory inulin and oligofuctose. The targets for their effects are the colonic microflora, the gastrointestinal physiology, the immune functions, the bioavailability of minerals, the metabolism of lipids and colonic carcinogenesis. Potential health benefits include reduction of risk of colonic diseases, noninsulin-dependent diabetes, obesity, osteoporosis and cancer. The documentation of such benefits requires scientific evidence that must be evaluated in terms of "health claims." Previous assessments have concluded that, in terms of "functional claims," strong evidence exists for a prebiotic effect and improved bowel habit. The evidence for calcium bioavailability is promising, and positive modulation of triglyceride metabolism is undergoing preliminary evaluation. Scientific research still must be done to support any "disease risk reduction claim," but sound hypotheses do already exist for designing the relevant human nutrition trials.

Biological Availability↗

Effects of dietary inulin on serum lipids.

Inulin is a carbohydrate belonging to a class of compounds known as fructans. Because inulin is resistant to digestion in the upper gastrointestional tract it reaches the large intestine essentially intact, where it is fermented by indigenous bacteria. Thus, it may be classified as a soluble dietary fiber. Soluble fibers have been shown to modulate serum lipids. A recent study examined the effect of consuming three servings per day of inulin-containing foods, compared with the effect of similar foods without inulin, on serum lipid profiles among hypercholesterolemic men and women. In addition, the practicality of including 18 g/d of inulin in a low fat diet was investigated. The recent study randomized, double-blind, crossover trial with two 6-wk treatment periods, separated by a 6-wk washout. Men and women (n = 21) with baseline LDL increased significantly (7.4 and 12.3%, respectively) during the control phase. There were small, nonsignificant declines in total (1.3%) and LDL-C (2.1%) during the inulin phase. Thus, differences in response between periods (inulin - control) were significant (P < 0.05) for LDL-C (-14.4%) and total cholesterol (-8.7%). Mild gastrointestinal discomfort was more common during the inulin than the control food phase; however, the gastrointestinal side-effect profile of inulin was similar to that of other soluble fibers. Although it was not possible to draw firm conclusions, inulin may have blunted the hypercholesterolemic effects observed during consumption of control foods. Additional research will be required to confirm the possible lipid-modulating properties of dietary inulin in humans.

Adult↗

The effect of synbiotics on colon carcinogenesis in rats.

Evidence indicates that consumption of probiotic microorganisms such as bifidobacteria reduces the risk of colon cancer in animal models. Feeding certain fructans such as oligofructose and inulin, which are thought to selectively increase the growth of intestinal bifidobacteria (i.e., a prebiotic effect), also has been shown to reduce colon cancer risk. The objective of our study was twofold, i. e., to determine whether the combination of bifidobacteria and oligofructose would have an additive effect (i.e., synbiotic) in reducing colon cancer risk in rats, and to determine whether other oligosaccharides would also be effective as part of a synbiotic combination. The development of colonic preneoplastic lesions (aberrant crypts) was used as an index of colon cancer risk. In one series of experiments, rats were given the carcinogen 1, 2-dimethylhydrazine (DMH) and administered one of the following treatments: skim milk (control), bifidobacteria (bifido), oligofructose (OF) or bifido + OF. Neither bifido nor OF alone significantly reduced aberrant crypt number. Bifido + OF reduced aberrant crypt number in five of six experiments, although the reduction was significant in only one. However, a paired comparison of the six experiments indicated a significant overall reduction in aberrant crypts by bifido + OF (P = 0.039). Soybean oligosaccharide (SBO) and wheat bran oligosaccharide (WBO) were also fed in combination with bifidobacteria. In two other experiments, SBO did not alter the number of aberrant crypts compared with the control, whereas WBO reduced aberrant crypt number in one experiment but not in another. Of OF, SBO and WBO, only SBO reduced the colonic mucosa proliferation compared with the control. These results suggest that the combination of bifidobacteria and oligofructose reduces colon cancer risk in carcinogen-treated rats, but the effect of other oligosaccharides is uncertain.

1,2-Dimethylhydrazine↗

Dietary oligofructose lessens hepatic steatosis, but does not prevent hypertriglyceridemia in obese zucker rats.

We studied the influence of oligofructose (OFS), a nondigestible fructan, on lipid metabolism in obese fa/fa Zucker rats. The addition of 10 g/100 g OFS to the diet slowed the increase in body weight without modifying serum triglycerides or glucose concentrations after 7 wk of treatment. However, an oral load of 2 g glucose and 5 g corn oil/kg body weight increased triglyceridemia more in OFS-fed rats than in control rats. After 10 wk, OFS decreased the hepatic concentration of triglycerides 57% relative to controls. The less severe steatosis was confirmed by histologic analysis. Among the key enzymes involved in fatty acid synthesis and esterification, only malic enzyme activity was significantly lower in OFS-fed rats than in controls. The epididymal fat mass was significantly lower in OFS-fed rats. In conclusion, dietary enrichment with OFS can counteract both the fat mass development and the hepatic steatosis that occur in obese Zucker rats. Future studies will be designed to clarify in obese animals the influence of dietary OFS on postprandial triglyceridemia, which is an important variable associated with the development of atherosclerosis in humans, and to analyze the biochemical mechanism underlying the "hepatoprotective" effect of OFS.

Analysis of Variance↗

Dietary chicory inulin increases whole-body bone mineral density in growing male rats.

Chicory inulin is a natural linear fructan that is not digested in the upper part of the gastrointestinal tract but is fermented in the cecocolon. It enhances calcium absorption in rats and improves femur and tibia mineral contents in gastrectomized or ovariectomized rats. We studied the effect of inulin (0, 5 and 10 g/100 g diet) on whole-body bone mineral content (WBBMC), whole-body bone area (WBBA) and whole-body bone mineral density (WBBMD) in live, growing male rats fed diets containing 0.2, 0.5 or 1 g Ca/100 g. Three experiments, each corresponding to one of the different dietary Ca concentrations, were performed using male Wistar rats (n = 108; 4 wk old). WBBMC was measured by dual-energy X-ray absorptiometry every 4 wk up to wk 22. Inulin increased WBBMC (P < 0.05) and WBBMD (P < 0.001) significantly but not WBBA at all ages and all dietary calcium concentrations. This is the first report to demonstrate that chicory inulin not only increases calcium absorption but also increases mineral parameters in whole-body bones.

Animals↗

Dietary inulin intake and age can affect intestinal absorption of zinc and copper in rats.

Nondigestible inulin-type fructan intake can stimulate intestinal mineral absorption in both humans and animals. However, this stimulatory effect may depend on experimental conditions such as the duration of the experience, mineral levels in the diet, and the animal's physiological status. The aim of this study was to determine the effect of inulin intake on Zn and Cu absorption in rats at different ages. Male Wistar rats (n = 80) of 4 different ages (2,5, 10, and 20 mo) were randomly assigned to a control group or a group administered 3.75% inulin in their diet for 4 d followed by 7.5% inulin for 26 d. Absorption of Zn67 and Cu65 was determined on d 21 of the experiment by fecal monitoring using Zn67 and Cu65 isotopes. Zn and Cu status was also assessed. Absorption of Zn67 and Cu65 was significantly lower in 11- and 21-mo-old rats than in 3- and 6 mo-old-rats. Moreover, inulin intake significantly increased Zn67 and Cu65 absorption. In conclusion, age and dietary inulin intake can significantly affect intestinal absorption of zinc and copper in rats. Further studies are required to explore this effect over longer periods of inulin intake and to test the effects of inulin in humans.

Aging↗

Pasture nonstructural carbohydrates and equine laminitis.

Fresh forages constitute a majority of the diet for many horses and ponies that graze on pastures during the growing season in many parts of the world. Grasses generally predominate in such pastures, with varying proportions of legumes. Nonstructural carbohydrates (NSC) (simple sugars, starch, and fructan) can induce laminitis experimentally, and NSC can accumulate to >400 g/kg of dry matter (DM) in pasture grasses. In this article we discuss the environmental factors affecting NSC accumulation in pastures and estimate the potential daily intakes of pasture NSC by grazing horses. We also discuss strategies for both reducing the NSC content of pastures and management practices that can help reduce intakes of pasture NSC by equines at risk of developing laminitis. This study reveals the importance of accurate forage analysis in the development of feeding regimens for equines at risk of laminitis.

Animal Feed↗

Redox regulation and storage processes during maturation in kernels of Triticum durum.

Metabolic changes during the development and maturation of Triticum durum Desf. (L.) kernels were studied, with particular emphasis on changes in the redox state of ascorbate and glutathione, as well as in the activities of the enzymes responsible for the recycling of their oxidized forms (ascorbic free radical reductase, EC 1.6.5.4; dehydroascorbate reductase, EC 1.8.5.1; glutathione reductase, EC 1.6.4.2) and for detoxification or utilization of hydrogen peroxide (ascorbate peroxidase, EC 1.11.1.11; catalase, EC 1.11.1.6). In parallel with this analysis, the production and storage of reserve compounds was studied, in particular, soluble carbohydrates (mono- di-saccharides and fructans) and the transition from sulphydryl groups to disulphide bridges into proteins. The results indicate that both the activities of the ascorbate and glutathione redox enzymes and that of catalase are high before the start of drying maturation, after which they decrease. Moreover, analysis of the redox state of ascorbate and glutathione pairs and the sulphydryl to disulphide transition into proteins suggests that these three parameters are tightly related during kernel maturation, thus confirming the involvement of the two redox pairs in protein maturation as well as in protection against reactive oxygen species. The physiological implications of the changes in cellular redox state and in soluble carbohydrates for the acquisition of desiccation tolerance and reaching the resting phase in orthodox seeds are also discussed.

Ascorbic Acid↗

Balancing supply and demand: the spatial regulation of carbon metabolism in grass and cereal leaves.

Leaf primary metabolism responds to changes in both supply of inputs and demand for products. Metabolic control in leaves changes both spatially and temporally. Using leaves of C(3) temperate Gramineae, the spatial control of carbohydrate metabolism has been studied using a range of approaches. Single-cell sampling and subsequent analysis of metabolites, proteins and transcripts has indicated significant differences between epidermal, mesophyll and parenchymatous bundle sheath cells. These differences correlate with differentiated function as heterotrophic, autotrophic and transport pathway components of the leaf. The review emphasizes the key role of sucrose and discusses its catabolism to hexoses and its anabolism to fructans as mechanisms for the preservation of sucrose gradients within the leaf.

Biological Transport↗

Oxygen deficiency affects carbohydrate reserves in overwintering forage crops.

Anaerobic conditions developing under an ice cover affect winter survival and spring regrowth of economically important perennial crops. The objective was to compare, during a prolonged period of low (<2%) O2 at low temperature, the concentration of carbohydrates of four plant species contrasting in their resistance to oxygen deficiency. Four perennial forage species, lucerne (Medicago sativa L.), red clover (Trifolium pratense L.), timothy (Phleum pratense L.), and cocksfoot (Dactylis glomerata L.) were subjected to a progressively developing oxygen deficiency stress by enclosing potted plants in gas-tight bags in late autumn for overwintering in an unheated greenhouse. Timothy was previously reported to be more resistant to oxygen deficiency than the three other species. Non-structural carbohydrates increased and remained at a higher concentration in timothy than in the other three species under low O2 concentration. Concentrations of sucrose, fructose, glucose, and fructans increased in response to oxygen deficiency in timothy, whereas the concentration of soluble sugars decreased under the same conditions in lucerne, red clover, and cocksfoot. The gene expression of glyceraldehyde-3-phosphate dehydrogenase increased in response to low oxygen concentration in oxygen deficiency-sensitive lucerne while it remained unchanged in the oxygen deficiency-resistant timothy. It is concluded that timothy maintains higher carbohydrate reserves under oxygen deficiency, a specific feature that could favour its winter survival and spring regrowth.

Carbohydrate Metabolism↗

Flower opening and closure: a review.

Flower opening and closure are traits of a reproductive syndrome, as it allows pollen removal and/or pollination. Various types of opening can be distinguished such as nocturnal and diurnal and single or repetitive. Opening is generally due to cell expansion. Osmotic solute levels increase by the conversion of polysaccharides (starch or fructan) to monosaccharides, and/or the uptake of sugars from the apoplast. Repeated opening and closure movements are often brought about by differential elongation. In tulip petals, for example, the upper and lower sides of the mesophyll exhibit a 10 degrees C difference in optimum temperature for elongation growth, resulting in opening in the morning and closure in the evening. Opening and closure in several other species is regulated by changes in light intensity and, in some species with nocturnal opening, by an increase in relative humidity. A minimum duration of darkness and light are usually required for opening and closure, respectively, in flowers that open during the day. Both phytochrome and a blue light receptor seem involved in light perception. In some species, opening and closure are regulated by an endogenous rhythm, which, in all cases investigated, can be reset by changes from dark to light and/or light to dark. So far, Arabidopsis mutants have not been used to investigate the timing of flower opening and closure. As its flowers open and close in a circadian fashion, several mutants that are involved in the circadian clock and its light input may help to provide an insight into this type of flower opening. The co-ordination of processes culminating in synchronized flower opening is, in many species, highly intricate. The complex control by endogenous and exogenous factors sets flower opening and closure apart from most other growth processes.

Circadian Rhythm↗

Cloning and characterization of a putative fructosyltransferase and two putative invertase genes from the temperate grass Lolium temulentum L.

The invertases of Lolium temulentum have been characterized at the enzyme level. However, studies on the expression of the genes coding for these enzymes have been lacking. To elucidate the role of acid 'invertase-like' genes in sucrose metabolism and carbon partitioning in Gramineae further, three 'invertase-like' homologous clones were isolated from L. temulentum cDNA expression libraries based on leaf tissue, using maize soluble invertase probes. The effect of developmental stage and alterations in carbohydrate status on the expression and tissue distribution of these genes was investigated. The three highly homologous genes (Inv 1:2, Inv 1:4, and FT 2:2) show different patterns of expression and different tissue distribution. Inv 1:2 was predominantly expressed in root tissue. Expression increased during the dark in root and tiller base tissue. Minimal variations in gene expression were observed in leaf tissue following changes in carbohydrate status. Inv 1:4 was predominantly expressed in tiller bases, leaf sheath, and leaf base, with increased expression in tissue samples in the dark period. FT 2:2 was also predominantly expressed in tiller bases, leaf sheath, and leaf base. Higher expression was observed in leaf tissue following increases in carbohydrate content, in a manner that paralleled the regulation and spatial occurrence of fructan in the leaf tissue. Whilst invertases and fructosyltransferases are difficult to distinguish at the level of the whole sequence, analysis of 5' sequence and specific amino acids allows discrimination which correlates with patterns of expression within the tissue. Based on expression patterns and sequence characteristics, it is proposed that Inv 1:2 and Inv 1:4 code for soluble acid invertases, whilst FT 2:2 codes for a fructosyltransferase.

Amino Acid Sequence↗

Identification of functionally important amino acid residues in Zymomonas mobilis levansucrase.

The catalytic residues of levansucrase (sucrose:2,6-beta-D-fructan 6-beta-D-fructosyltransferase, EC 2.4.1.10) from Zymomonas mobilis were analyzed by random mutation and site-directed mutagenesis. We found that substitution of Glu278 with Asp and His reduced the k(cat) for sucrose hydrolysis 30- and 210-fold, respectively, strongly suggesting Glu278 plays a key role in catalyzing this reaction. Given the likelihood that another acidic amino residue was also involved, we constructed variants in which acidic amino acids located within homologous regions among bacterial levansucrases and fructosyltransferases were substituted, and found that substitution of Asp194, located in homologous region III, abolished sucrose hydrolysis. In addition, Glu278 was determined to be situated within the DXXER motif in homologous region IV conserved among bacterial levansucrases and fructosyltransferases, while Asp194 was within the triplet RDP motif conserved among bacterial levansucrases, fructosyltransferases and fructofuranosidases. Finally, comparison of our findings with published data on other site-directed mutated enzymes indicated His296, also located in homologous region IV, is crucial for catalysis of the transfructosylation reaction.

Amino Acid Sequence↗