Acetylation of homogalacturonan and rhamnogalacturonan-I is catalyzed by a suite of trichome birefringence-like proteins.

Plant cell wall polysaccharides, including xylan, mannan, xyloglucan, and pectins, are often acetylated and members of the domain of unknown function 231 (DUF231)/trichome birefringence-like (TBL) family have been shown to be O-acetyltransferases mediating the acetylation of xylan, mannan, and xyloglucan. However, little is known about the O-acetyltransferases responsible for pectin acetylation. In this report, we biochemically characterized a suite of Arabidopsis DUF231/TBL proteins for their roles in pectin acetylation. We generated 24 TBL recombinant proteins in mammalian cells and demonstrated that 10 of them were able to transfer acetyl groups from acetyl-CoA onto the pectins homogalacturonan (HG) or rhamnogalacturonan-I (RG-I), and thus were named pectin O-acetyltransferase 1 to 10 (POAT1 to 10). It was found that POAT2,4,9,10 specifically acetylated HG and POAT5,6 acetylated RG-I, whereas POAT1,3,7,8 could act on both HG and RG-I. The acetylation of HG and RG-I by POATs was further corroborated by hydrolysis with pectin acetylesterases and by nuclear magnetic resonance spectroscopy. In addition, mutations of the conserved GDS and DXXH motifs in POAT3 and POAT8 were shown to lead to a loss of their ability to acetylate HG and RG-I. Furthermore, simultaneous RNA interference downregulation of POAT1,3,6,7,8 resulted in reduced cell expansion, impaired plant growth, and decreased pectin acetylation. Together, our findings indicate that these POATs are pectin O-acetyltransferases involved in acetylation of the pectin polysaccharides HG and RG-I.

Identification of intracellular activation mechanism of rhamnogalacturonan-I type polysaccharide purified from Panax ginseng leaves in macrophages and roles of component sugar chains on activity.

This study aimed to investigate the mechanisms underlying intracellular signaling pathways in macrophages in relation to the structural features of rhamnogalacturonan (RG) I-type polysaccharide (PGEP-I) purified from Panax ginseng leaves. For this investigation, we used several specific inhibitors and antibodies against mitogen-activated protein kinase (MAPK), nuclear factor-kappa B (NF-κB), and pattern recognition receptors (PRRs). Furthermore, we investigated the roles of component sugar chains on immunostimulating activity through a sequential enzymatic and chemical degradation steps. We found that PGEP-I effectively induced the phosphorylation of several MAPK- and NF-κB-related proteins, such as p38, cJun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and p65. Particularly, immunocytochemistry analysis confirmed the PGEP-I-induced translocation of p65 into the nucleus. Furthermore, the breakdown of PGEP-I side chains and main chain during sequential enzymatic and chemical degradation reduced the PGEP-I-induced macrophage cytokine secretion activity. IL-6, TNF-α, and NO secreted by macrophages are associated with several signaling pathway proteins such as ERK, JNK, and NF-κB and several PRRs such as dectin-1, CD11b, CD14, TLR2, TLR4, and SR. Thus, these findings suggest that PGEP-I exerts potent macrophage-activating effects, which can be attributed to its typical RG-I structure comprising arabinan, type II arabinogalactan, and rhamnose-galacturonic acid repeating units in the main chain.

Suppressing the rhamnogalacturonan lyase gene FaRGLyase1 preserves RGI pectin degradation and enhances strawberry fruit firmness.

Plant rhamnogalacturonan lyases (RGLyases) cleave the backbone of rhamnogalacturonan I (RGI), the "hairy" pectin and polymer of the disaccharide rhamnose (Rha)-galacturonic acid (GalA) with arabinan, galactan or arabinogalactan side chains. It has been suggested that RGLyases could participate in remodeling cell walls during fruit softening, but clear evidence has not been reported. To investigate the role of RGLyases in strawberry softening, a genome-wide analysis of RGLyase genes in the genus Fragaria was performed. Seventeen genes encoding RGLyases with functional domains were identified in Fragaria × ananassa. FaRGLyase1 was the most expressed in the ripe receptacle of cv. Chandler. Transgenic strawberry plants expressing an RNAi sequence of FaRGLyase1 were obtained. Three transgenic lines yielded ripe fruits firmer than controls without other fruit quality parameters being significantly affected. The highest increase in firmness achieved was close to 32%. Cell walls were isolated from ripe fruits of two selected lines. The amount of water-soluble and chelated pectins was higher in transgenic lines than in the control. A carbohydrate microarray study showed a higher abundance of RGI epitopes in pectin fractions and in the cellulose-enriched fraction obtained from transgenic lines. Sixty-seven genes were differentially expressed in transgenic ripe fruits when compared with controls. These genes were involved in various physiological processes, including cell wall remodeling, ion homeostasis, lipid metabolism, protein degradation, stress response, and defense. The transcriptomic changes observed in FaRGLyase1 plants suggest that senescence was delayed in transgenic fruits.

Recovering rhamnogalacturonan-I pectin from sugar beet pulp using a sequential ultrasound and microwave-assisted extraction: Study on extraction optimization and membrane purification.

This research focuses on the integrated recovery of rhamnogalacturonan-I (RG-I) pectin from sugar beet pulp (SBP). First, the extraction of RG-I pectin through sequential ultrasound-assisted extraction (UAE) and microwave-assisted extraction (MAE) was assessed. Optimization using a response surface methodology identified the optimal conditions as initial pH 4, 10 min of UAE, and 157 °C for MAE, achieving a 66.0 % recovery of pectooligosaccharides (POS). Additionally, purification through continuous diafiltration and concentration via ultrafiltration of the POS using membranes with different molecular weight cut-offs (MWCO) was explored. In contrast to previous research using discontinuous diafiltration, the use of continuous diafiltration allowed a decrease in the extract viscosity and obtained higher yields using a higher MWCO membrane. The refined RG-I pectin solids exhibited a high global yield (39-40 g pectin/100 g SBP), and high-methoxyl characteristics, as well as purity levels (70-80 %) similar to commercial prebiotics.

Grape-Derived Polysaccharide Extracts Rich in Rhamnogalacturonans-II as Potential Modulators of White Wine Flavor Compounds.

Many authors have investigated the role of mannoproteins on wine quality, but very few have analyzed the use of grape-derived polysaccharides as they are not commercially available. In this study, purified grape-derived polysaccharides from red wine (WPP) and winemaking by-products (DWRP: Distilled Washing Residues Polysaccharides) were used as potential fining agents to modulate white wine flavor. Phenolics and volatile compounds were analyzed in the control and wines treated with WPP, DWRP, and commercial mannoproteins (CMs) after one and twelve months of bottling, and a sensory analysis was conducted. WPP and DWRP, rich in rhamnogalacturonans-II, showed themselves to be good modulators of wine aroma and astringency. Improvement in wine aroma was related to an increase in all volatile families expect higher alcohols and volatile acids. The modulation of astringency and bitterness was related to a reduction in the proanthocyanidin content and its mean degree of polymerization. Extracts with polysaccharides with higher protein contents presented a higher retention of volatile compounds, and DWRP extract had more positive effects on the overall aroma. Our novel results present the possibility of obtaining valuable polysaccharides from distilled washing residues of wine pomaces, which could promote its valorization as a by-product. This is the first time the potential use of this by-product has been described.

Cellulose assembles into helical bundles of uniform handedness in cell walls with abnormal pectin composition.

Plant cells and organs grow into a remarkable diversity of shapes, as directed by cell walls composed primarily of polysaccharides such as cellulose and multiple structurally distinct pectins. The properties of the cell wall that allow for precise control of morphogenesis are distinct from those of the individual polysaccharide components. For example, cellulose, the primary determinant of cell morphology, is a chiral macromolecule that can self-assemble in vitro into larger-scale structures of consistent chirality, and yet most plant cells do not display consistent chirality in their growth. One interesting exception is the Arabidopsis thaliana rhm1 mutant, which has decreased levels of the pectin rhamnogalacturonan-I and causes conical petal epidermal cells to grow with a left-handed helical twist. Here, we show that in rhm1 the cellulose is bundled into large macrofibrils, unlike the evenly distributed microfibrils of the wild type. This cellulose bundling becomes increasingly severe over time, consistent with cellulose being synthesized normally and then self-associating into macrofibrils. We also show that in the wild type, cellulose is oriented transversely, whereas in rhm1 mutants, the cellulose forms right-handed helices that can account for the helical morphology of the petal cells. Our results indicate that when the composition of pectin is altered, cellulose can form cellular-scale chiral structures in vivo, analogous to the helicoids formed in vitro by cellulose nano-crystals. We propose that an important emergent property of the interplay between rhamnogalacturonan-I and cellulose is to permit the assembly of nonbundled cellulose structures, providing plants flexibility to orient cellulose and direct morphogenesis.

Preparation of branched RG-I-rich pectin from red dragon fruit peel and the characterization of its probiotic properties.

Red dragon fruit peel is a pectin-rich fruit waste that is a potential source of prebiotics and whose different sources and structures will influence its prebiotic function. Thus, we compared the effects of three extraction methods on the structure and prebiotic function of red dragon fruit pectin, the results showed that the citric acid extracted pectin produced a high Rhamnogalacturonan-I (RG-I) region (66.59 mol%) and more side-chains of Rhamnogalacturonan-I ((Ara + Gal)/Rha = 1.25), which can promote bacterial proliferation significantly. The side-chains of Rhamnogalacturonan-I may be an important factor in that pectin can promote the proliferation of B. animalis. Our results provide a theoretical basis for the prebiotic application of red dragon fruit peel.

Time-dependent fermentation of different structural units of commercial pectins with intestinal bacteria.

Many of the proposed health-related properties of pectins are based on their fermentability in the large intestine, but detailed structure-related studies on pectin fermentation have not been reported so far. Here, pectin fermentation kinetics were studied with a focus on structurally different pectic polymers. Therefore, six commercial pectins from citrus, apple, and sugar beet were chemically characterized and fermented in in vitro fermentation assays with human fecal samples over different periods of time (0 h, 4 h, 24 h, 48 h). Structure elucidation of intermediate cleavage products showed differences in fermentation speed and/or fermentation rate among the pectins, but the order in which specific structural pectic elements were fermented was comparable across all pectins. Neutral side chains of rhamnogalacturonan type I were fermented first (between 0 and 4 h), followed by homogalacturonan units (between 0 and 24 h) and, at last, the rhamnogalacturonan type I backbone (between 4 and 48 h). This indicates that fermentation of different pectic structural units might take place in different sections of the colon, potentially affecting their nutritional properties. For the formation of different short-chain fatty acids, mainly acetate, propionate, and butyrate, and the influence on microbiota, there was no time-dependent correlation regarding the pectic subunits. However, an increase of members of the bacterial genera Faecalibacterium, Lachnoclostridium, and Lachnospira was observed for all pectins.

Making and breaking of boron bridges in the pectic domain rhamnogalacturonan-II at apoplastic pH in vivo and in vitro.

Cross-linking of the cell-wall pectin domain rhamnogalacturonan-II (RG-II) via boron bridges between apiose residues is essential for normal plant growth and development, but little is known about its mechanism or reversibility. We characterized the making and breaking of boron bridges in vivo and in vitro at 'apoplastic' pH. RG-II (13-26 µm) was incubated in living Rosa cell cultures and cell-free media with and without 1.2 mm H3 BO3 and cationic chaperones (Ca2+ , Pb2+ , polyhistidine, or arabinogalactan-protein oligopeptides). The cross-linking status of RG-II was monitored electrophoretically. Dimeric RG-II was stable at pH 2.0-7.0 in vivo and in vitro. In-vitro dimerization required a 'catalytic' cation at all pHs tested (1.75-7.0); thus, merely neutralizing the negative charge of RG-II (at pH 1.75) does not enable boron bridging. Pb2+ (20-2500 µm) was highly effective at pH 1.75-4.0, but not 4.75-7.0. Cationic peptides were effective at approximately 1-30 µm; higher concentrations caused less dimerization, probably because two RG-IIs then rarely bonded to the same peptide molecule. Peptides were ineffective at pH 1.75, their pH optimum being 2.5-4.75. d-Apiose (>40 mm) blocked RG-II dimerization in vitro, but did not cleave existing boron bridges. Rosa cells did not take up d-[U-14 C]apiose; therefore, exogenous apiose would block only apoplastic RG-II dimerization in vivo. In conclusion, apoplastic pH neither broke boron bridges nor prevented their formation. Thus boron-starved cells cannot salvage boron from RG-II, and 'acid growth' is not achieved by pH-dependent monomerization of RG-II. Divalent metals and cationic peptides catalyse RG-II dimerization via co-ordinate and ionic bonding respectively (possible and impossible, respectively, at pH 1.75). Exogenous apiose may be useful to distinguish intra- and extra-protoplasmic dimerization.

Most of the rhamnogalacturonan-I from cultured Arabidopsis cell walls is covalently linked to arabinogalactan-protein.

To characterize a purified rhamnogalacturonan-I (RG-I) containing both RG-I and arabinogalactan-protein (AGP) types of glycosyl residues, an AGP-specific ß-1,3-galactanase that can cleave the AG backbone and release the AG sidechain was applied to this material. Carbohydrate analysis and NMR spectroscopy verified that the galactanase-released carbohydrate consists of RG-I covalently attached to the AG sidechain, proving a covalent linkage between RG-I and AGP. Size exclusion chromatography-multiangle light scattering-refractive index detection revealed that the galactanase-released RG-I has an average molecular weight of 41.6 kDa, which, together with the percentage of pectic sugars suggests an RG-I-AGP comprising one AGP covalently linked to two RG-I glycans. Carbohydrate analysis and NMR results of the RG-I-AGP, the galactanase-released glycans, and the RG lyase-released glycans demonstrated that the attached RG-I glycans are decorated with α-1,5-arabinan, ß-1,4-galactan, xylose, and 4-O-Me-xylose sidechains. Our measurement suggests that the covalently linked RG-I-AGP is the major component of the traditionally prepared RG-I.

Rhamnogalacturonan-I enriched pectin from steamed ginseng ameliorates lipid metabolism in type 2 diabetic rats via gut microbiota and AMPK pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax ginseng C. A. Meyer (Ginseng) has traditionally been used to treat diabetes. Polysaccharide is the main active component of ginseng, and has been proved to have hypoglycaemic and hypolipidaemic effects, but its mechanism remains unclear. AIM OF THE STUDY: This study aimed to evaluate the effect and the potential mechanism of rhamnogalacturonan-I enriched pectin (GPS-1) from steamed ginseng on lipid metabolism in type 2 diabetes mellitus (T2DM) rats. MATERIALS AND METHODS: GPS-1 was prepared by water extraction, ion-exchange and gel chromatography. High-glucose/high-fat diet combined with streptozotocin was used to establish T2DM rat models, and lipid levels in serum and liver were tested. 16S rRNA sequencing and gas chromatography-mass spectrometry were used to detect the changes of gut microbiota and metabolites. The protein and mRNA levels of lipid synthesis-related genes were detected by Western blot and quantitative real-time polymerase chain reaction. RESULTS: The polyphagia, polydipsia, weight loss, hyperglycaemia, hyperlipidaemia and hepatic lipid accumulation in T2DM rats were alleviated after GPS-1 intervention. GPS-1 modulated the gut microbiota composition of T2DM rats, increased the levels of short-chain fatty acids, and promoted the secretion of glucagon-like peptide-1 and peptide tyrosine tyrosine. Further, GPS-1 activated AMP-activated protein kinases, phosphorylated acetyl-CoA carboxylase, reduced the expression of sterol regulatory element-binding protein-1c and fatty acid synthases in T2DM rats. CONCLUSIONS: The regulation effects of GPS-1 on lipid metabolism in T2DM rats are related to the regulation of gut microbiota and activation of AMP-activated protein kinase pathway.

Structural Characteristics of Polysaccharide GP2a in Gardenia jasminoides and Its Immunomodulatory Effect on Macrophages.

Here, we elucidated the structural characteristics of a polysaccharide isolated from Gardenia jasminoides Ellis (labeled as GP2a) and its immunomodulatory activity. GP2a is an acidic polysaccharide with a molecular weight of 44.8 kDa, mostly comprising galacturonic acid. Methylation analysis revealed 4-GalpA (74.8%) to be the major sugar residue in GP2a. Nuclear magnetic resonance analysis indicated that its main chain comprised →4)-α-D-GalpA-6-OMe-(1→4)-α-D-GalpA-(1→ and →4)-α-D-GalpA-6-OMe-(1→2)-α-L-Rhap-(1→, with galactan and arabinans linked to the C-4 position of →2)-α-L-Rhap-(1→ residue as branched chains. Furthermore, GP2a showed no obvious toxicity to macrophages (RAW 264.7) while enhancing cell viability in a dose- and time-dependent manner. Compared with untreated cells, nitric oxide production and secretion of cytokines, such as tumor necrosis factor-α, interferon-γ, interleukin (IL)-1ß, IL-6, and granulocyte macrophage colony stimulating factor, in GP2a-treated cells significantly increased after 48 h. At 300 µg/mL GP2a concentration, there was no significant difference in the cytokine levels in GP2a- and lipopolysaccharide-treated cells (the positive control). In summary, GP2a is a pectic polysaccharide with homogalacturonan and rhamnogalacturonan-I structural regions in the main chain. Based on its immunomodulatory effects in vitro, GP2a may have potential uses in functional food and medicine.

Structural characteristic of pectin-glucuronoxylan complex from Dolichos lablab L. hull.

Polysaccharides are important constituents in Dolichos lablab hull. Herein, pectin-glucuronoxylan complex from D. lablab hull designated as DLHP-3 (D. lablab hull polysaccharide,) was prepared by ion exchange and gel permeation chromatography, and further characterized by acid degradation and enzymatic hydrolysis, methylation combined with GC-MS, NMR and MALDI-TOF-MS analysis. Both of pectin and glucuronoxylan regions were found in DLHP-3. The glucuronoxylan region consisted of a →4)-ß-Xylp-(1→ backbone with branches of α-GlcpA-(1→ substituted at O-2 site, and the ratio of xylose to glucuronic acid was about 5:1. Acetyl groups were mainly attached to O-3 site of →2,4)-ß-Xylp-(1→ residues. The main chain of pectin region could be represented by →4)-α-GalpA-(1→4)-α-GalpA-(1→ and →2)-α-Rhap-(1→4)-α-GalpA-(1→ with partial methyl-esterification. The side chains were deduced to embrace arabinan and arabinogalactan linked to rhamnogalacturonan-I region. Pectin was probably covalently bound to glucuronoxylan. Our findings uncovered the molecular structure of pectin-glucuronoxylan complex from D. lablab hull.

The potential of subcritical water as a "green" method for the extraction and modification of pectin: A critical review.

Subcritical water is a "green" method of extraction and modification of pectin being explored in recent times. While the conventional acid extraction degrades the side chains and produces homogalacturonan (HG)-rich pectic polysaccharides, subcritical water extraction preserved the hairy region, namely the rhamnogalacturonan-I (RG-I) region of the pectin. However, higher temperatures (usually greater than 160 °C) degraded the RG-I and HG motifs, producing pectic oligosaccharides. A high selectivity towards pectic polysaccharides with a low protein content was observed during extraction by subcritical water. This can be majorly attributed to the heat-induced denaturation of proteins. Although the bioactive and emulsifying properties were more remarkable for subcritical water-extracted pectin, the rheological properties such as elasticity were negatively impacted. Apart from extraction, subcritical water can also be employed to aid the breakdown of pectic polysaccharides into oligosaccharides. The addition of several organic acids in subcritical water can help form pectic fragments, which are otherwise possible only by adding a cocktail of enzymes. For instance, carboxylic acids in subcritical water media can have a similar action to endo-polygalacturonase on the homogalacturonan backbone. It is worthwhile to note that intense extraction or modification conditions can form advanced glycation end products, which are undesirable and should be monitored throughout the modification process. Several thermodynamic and kinetic models can be employed to predict the breakdown of the pectin structure in subcritical conditions. Finally, this study suggests a strategy for obtaining the optimum process parameters, namely, temperature, duration, and the liquid:solid ratio for achieving maximum yield and the desired structure of the pectic polysaccharide.

Effects of Dietary Supplementation with Carrot-Derived Rhamnogalacturonan-I (cRG-I) on Accelerated Protective Immune Responses and Quality of Life in Healthy Volunteers Challenged with Rhinovirus in a Randomized Trial.

An adequate and balanced supply of nutrients is essential for maintaining health, and an optimal immune response is fast, contained and properly controlled, curbing infections quickly while minimizing damage. Several micronutrients contribute to normal immune function and certain dietary fibers, for example pectic polysaccharides, can play an important role in educating and regulating immune cell responses. The aim of this paper is to elaborate on our initial findings that dietary supplementation with carrot-derived rhamnogalacturonan-I (cRG-I) accelerates and augments local innate immune and anti-viral interferon response to a rhinovirus-16 (RV16) infection and reduces the severity and duration of symptoms in humans. Dietary intake of cRG-I also enhanced immune responses to this respiratory viral infection as measured by ex vivo stimulation of whole blood with the Toll-like receptor 3 (TLR3) ligand polyinosinic:polycytidylic acid and NK cell function. Consumption of cRG-I also reduced the negative effects of this common cold infection on quality of life as assessed by individual symptom scores. RG-I from carrot is a safe, sustainable, and economically viable solution that could easily be integrated into food products and dietary supplements aiming to support immune fitness and wellbeing.

An Arabidopsis thaliana arabinogalactan-protein (AGP31) and several cationic AGP fragments catalyse the boron bridging of rhamnogalacturonan-II.

Rhamnogalacturonan-II (RG-II) is a complex pectic domain in plant primary cell walls. In vivo, most RG-II domains are covalently dimerised via borate diester bridges, essential for correct cell-wall assembly, but the dimerisation of pure RG-II monomers by boric acid in vitro is extremely slow. Cationic 'chaperones' can promote dimerisation, probably by overcoming the mutual repulsion between neighbouring anionic RG-II molecules. Highly effective artificial chaperones include Pb2+ and polyhistidine, but the proposed natural chaperones remained elusive. We have now tested cationic peptide fragments of several Arabidopsis thaliana arabinogalactan-proteins (AGPs) as candidates. Fragments of AGP17, 18, 19 and 31 were effective, typically at ∼25 µg/ml (9-19 µM), promoting the boron bridging of 16-20 µM monomeric RG-II at pH 4.8 in vitro. Native AGP31 glycoprotein was also effective, and hexahistidine was moderately so. All chaperones tested interacted reversibly with RG-II and were not consumed during the reaction; thus they acted catalytically, and may constitute the first reported boron-acting enzyme activity, an RG-II borate diesterase. Many of the peptide chaperones became less effective catalysts at higher concentration, which we interpret as due to the formation of RG-II-peptide complexes with a net positive charge, as mutually repulsive as negatively charged pure RG-II molecules. The four unique AGPs studied here may serve an enzymic role in the living plant cell, acting on RG-II within Golgi cisternae and/or in the apoplast after secretion. In this way, RG-II and specific AGPs may contribute to cell-wall assembly and hence plant cell expansion and development.

Boron bridging of rhamnogalacturonan-II in Rosa and arabidopsis cell cultures occurs mainly in the endo-membrane system and continues at a reduced rate after secretion.

BACKGROUND AND AIMS: Rhamnogalacturonan-II (RG-II) is a domain of primary cell-wall pectin. Pairs of RG-II domains are covalently cross-linked via borate diester bridges, necessary for normal cell growth. Interpreting the precise mechanism and roles of boron bridging is difficult because there are conflicting hypotheses as to whether bridging occurs mainly within the Golgi system, concurrently with secretion or within the cell wall. We therefore explored the kinetics of RG-II bridging. METHODS: Cell-suspension cultures of Rosa and arabidopsis were pulse-radiolabelled with [14C]glucose, then the boron bridging status of newly synthesized [14C]RG-II domains was tracked by polyacrylamide gel electrophoresis of endo-polygalacturonase digests. KEY RESULTS: Optimal culture ages for 14C-labelling were ~5 and ~1 d in Rosa and arabidopsis respectively. De-novo [14C]polysaccharide production occurred for the first ~90 min; thereafter the radiolabelled molecules were tracked as they 'aged' in the wall. Monomeric and (boron-bridged) dimeric [14C]RG-II domains appeared simultaneously, both being detectable within 4 min of [14C]glucose feeding, i.e. well before the secretion of newly synthesized [14C]polysaccharides into the apoplast at ~15-20 min. The [14C]dimer : [14C]monomer ratio of RG-II remained approximately constant from 4 to 120 min, indicating that boron bridging was occurring within the Golgi system during polysaccharide biosynthesis. However, [14C]dimers increased slightly over the following 15 h, indicating that limited boron bridging was continuing after secretion. CONCLUSIONS: The results show where in the cell (and thus when in the 'career' of an RG-II domain) boron bridging occurs, helping to define the possible biological roles of RG-II dimerization and the probable localization of boron-donating glycoproteins or glycolipids.

Tumor inhibitory effect via immunostimulating activities of a rhamnogalacturonan-I-rich polysaccharide isolated from turmeric (Curcuma longa L.).

In this study, a turmeric polysaccharide (TP-0) was isolated through hot water extraction and ethanol precipitation to produce a novel active polysaccharide from turmeric other than curcuminoids. TP-0 was found to be primarily composed of eight different monosaccharides, such as galactose (15.9%), galacturonic acid (15.2%), arabinose (11.4%), and rhamnose (9.7%), which are typical rhamnogalacturonan (RG)-I sugars. When stimulated with TP-0, peritoneal macrophages secreted a variety of immunostimulatory cytokines. In addition, intravenous and oral administration of TP-0 significantly enhanced the natural killer (NK) cells and cytotoxic T lymphocyte (CTL)-mediated cytotoxicity against tumor cells. In an assay for lung cancer induced by Colon26-M3.1 carcinoma, prophylactic intravenous and oral administration of TP-0 effectively inhibited lung cancer. These findings reveal that TP-0, a typical RG-I-type polysaccharide that is isolated from turmeric, has potent anti-metastatic activities, and these activities are linked to various immunological factors such as macrophages, NK cells, and CTL. PRACTICAL APPLICATIONS: Many studies related with turmeric have only focused that a curcuminoid of turmeric has beneficial effects on human health system. Nevertheless, in this study, it was confirmed that polysaccharide isolated from turmeric showed potent anti-cancer effects via activities of various immunological factors such as macrophages, NK cells, and CTL. These results suggest the high potential for development value of turmeric as a new candidate for immunostimulating-related health functional food ingredients.

Immunostimulating and intracellular signaling pathways mechanism on macrophage of rhamnogalacturonan-I type polysaccharide purified from radish leaves.

In this study, the intracellular signaling pathways involved in macrophage activation through the RG-I-type polysaccharide (REP-I) purified from radish leaves were elucidated. The gene expression and secretion of immune-related factors such as interleukin (IL)-6, tumor necrosis factor (TNF)-α, and nitrogen oxide (NO) from macrophages were enhanced by the addition of REP-I. Moreover, immunoblotting and immunocytochemistry analyses indicated that REP-I dose-dependently phosphorylated the mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) pathways. An investigation using different inhibitors revealed that the effect of REP-I on NO secretion was mostly promoted by c-Jun N-terminal kinase (JNK) and NF-κB. Furthermore, the secretion of IL-6 was mostly induced via extracellular-signal-regulated kinase (ERK), JNK, and NF-κB. TNF-α secretion was mostly induced via NF-κB. In contrast, an investigation using anti-pattern recognition receptor (PRR) antibodies revealed that the effect of REP-I on the secretion of NO was mostly related with dectin-1, scavenger receptor (SR), toll-like receptor (TLR)2, TLR4, CD14, and CD11b. Furthermore, the secretion of IL-6 was mostly involved with SR, and the secretion of TNF-α was mostly relevance to TLR2. In conclusion, it is affirmed that immunostimulatory activation of macrophage of REP-I purified from radish leaves was deeply associated with several PRR and phosphorylating MAPK and NF-κB.