Retracing Storage Polysaccharide Evolution in Stramenopila

Eukaryotes most often synthesize storage polysaccharides in the cytosol or vacuoles in the form of either alpha (glycogen/starch)- or beta-glucosidic (chrysolaminarins and paramylon) linked glucan polymers. In both cases, the glucose can be packed either in water-soluble (glycogen and chrysolaminarins) or solid crystalline (starch and paramylon) forms with different impacts, respectively, on the osmotic pressure, the glucose accessibility, and the amounts stored. Glycogen or starch accumulation appears universal in all free-living unikonts (metazoa, fungi, amoebozoa, etc.), as well as Archaeplastida and alveolata, while other lineages offer a more complex picture featuring both alpha- and beta-glucan accumulators. We now infer the distribution of these polymers in stramenopiles through the bioinformatic detection of their suspected metabolic pathways. Detailed phylogenetic analysis of key enzymes of these pathways correlated to the phylogeny of Stramenopila enables us to retrace the evolution of storage polysaccharide metabolism in this diverse group of organisms. The possible ancestral nature of glycogen metabolism in eukaryotes and the underlying source of its replacement by beta-glucans are discussed.

Euglena Gracilis and β-Glucan Paramylon Induce Ca2+ Signaling in Intestinal Tract Epithelial, Immune, and Neural Cells

The intestinal tract contains over half of all immune cells and peripheral nerves and manages the beneficial interactions between food compounds and the host. Paramylon is a β-1,3-glucan storage polysaccharide from Euglena gracilis (Euglena) that exerts immunostimulatory activities by affecting cytokine production. This study investigated the signaling mechanisms that regulate the beneficial interactions between food compounds and the intestinal tract using cell type-specific calcium (Ca2+) imaging in vivo and in vitro. We successfully visualized Euglena- and paramylon-mediated Ca2+ signaling in vivo in intestinal epithelial cells from mice ubiquitously expressing the Yellow Cameleon 3.60 (YC3.60) Ca2+ biosensor. Moreover, in vivo Ca2+ imaging demonstrated that the intraperitoneal injection of both Euglena and paramylon stimulated dendritic cells (DCs) in Peyer’s patches, indicating that paramylon is an active component of Euglena that affects the immune system. In addition, in vitro Ca2+ imaging in dorsal root ganglia indicated that Euglena, but not paramylon, triggers Ca2+ signaling in the sensory nervous system innervating the intestine. Thus, this study is the first to successfully visualize the direct effect of β-1,3-glucan on DCs in vivo and will help elucidate the mechanisms via which Euglena and paramylon exert various effects in the intestinal tract.

Production of Insoluble Starch-Like Granules in Escherichia coli by Modification of the Glycogen Synthesis Pathway

While investigating the conversion of cellulosic biomass to starch-like materials for industrial use, it was observed that the overexpression of native ADP-glucose pyrophosphorylase GlgC in Escherichia coli led to the formation of insoluble polysaccharide granules within the cytoplasm, occupying a large fraction of the cell volume, as well as causing an overall increase in cellular polysaccharide content. TEM microscopy revealed that the granules did not have the lamellar structure of starch, but rather an irregular, clustered structure. On starvation, cells overexpressing GlgC appeared unable to fully degrade their polysaccharide material and granules were still clearly visible in cultures after 8 days of starvation. Interestingly, the additional overexpression of the branching enzyme GlgB eliminated the production of granules and led to a further increase in cellular polysaccharides. GlgC is generally thought to be responsible for the rate-limiting step of glycogen synthesis. Our interpretation of these results is that excess GlgC activity may cause the elongation of glycogen chains to outpace the addition of side branches, allowing the chains of adjacent glycogen molecules to reach lengths at which they spontaneously intertwine, forming dense clusters that are largely inaccessible to the host. However, upon additional upregulation of the GlgB branching enzyme, the branching of the polysaccharide is able to keep speed with the synthesis of linear chains, eliminating the granule phenotype. This study suggests potential avenues for increasing bacterial polysaccharide production and recovery.ImportanceIn this work, the polysaccharide stores of Escherichia coli were altered through the addition of extra copies of the bacteria’s own polysaccharide synthesis genes. In this way, bacteria were created that produced over twice the level of storage polysaccharide as a control strain, in the form of a granule that could potentially facilitate easy harvest. Another form of mutant Escherichia coli was created that produced over seven times the normal level of storage polysaccharide, and also grew to higher cell densities in liquid culture. In addition to increasing our understanding of glycogen synthesis, it is proposed that similarly modified bacteria, grown on inexpensive waste materials, may be a useful source of starch-like polysaccharides for industrial or agricultural use. In particular, the use of cyanobacterial glycogen as a carbon source for biofuels has recently been gaining interest, and the work presented here may well be applicable in this field.

Compositional and Structural Features of the Main Bioactive Polysaccharides Present in the Aloe vera Plant

Aloe vera (A. barbadensis Miller) is probably one of the most popular plants, widely studied because of numerous properties associated with the polysaccharides present in its gel. In particular, two main types of bioactive polysaccharides can be distinguished in the A. vera gel: an acetylated mannose-rich polymer that functions as storage polysaccharide, and a galacturonic acid–rich polymer as the main component comprising the cell walls of the parenchymatous tissue. Interestingly, most of the beneficial properties related to the aloe plant have been associated with the acetylated mannose-rich polysaccharide, also known as acemannan. However, the composition and structural features of these polysaccharides, as well as the beneficial properties associated with them, may be altered by different factors, such as the climate, soil, postharvest treatments, and processing. Further, different analytical methods have been used not only to identify but also to characterize the main polysaccharides found in parenchyma of A. vera leaf. Within this context, the main aim of this review is to summarize the most relevant information about the structural and compositional features of the main polysaccharides found in the A. vera gel as well as the most relevant analytical techniques used for their identification and their influence on the technological, functional, and beneficial properties related to the A. vera plant.

Structural and biochemical characterization of the laminarinaseZgLamCGH16fromZobellia galactanivoranssuggests preferred recognition of branched laminarin

Laminarin is a β-1,3-D-glucan displaying occasional β-1,6 branches. This storage polysaccharide of brown algae constitutes an abundant source of carbon for marine bacteria such asZobellia galactanivorans. This marine member of the Bacteroidetes possesses five putative β-1,3-glucanases [four belonging to glycosyl hydrolase family 16 (GH16) and one to GH64] with various modular architectures. Here, the characterization of the β-glucanaseZgLamC is reported. The catalytic GH16 module (ZgLamCGH16) was produced inEscherichia coliand purified. This recombinant enzyme has a preferential specificity for laminarin but also a significant activity on mixed-linked glucan (MLG). The structure of an inactive mutant ofZgLamCGH16in complex with a thio-β-1,3-hexaglucan substrate unravelled a straight active-site cleft with three additional pockets flanking subsites −1, −2 and −3. These lateral pockets are occupied by a glycerol, an acetate ion and a chloride ion, respectively. The presence of these molecules in the vicinity of the O6 hydroxyl group of each glucose moiety suggests thatZgLamCGH16accommodates branched laminarins as substrates. Altogether,ZgLamC is a secreted laminarinase that is likely to be involved in the initial step of degradation of branched laminarin, while the previously characterizedZgLamA efficiently degrades unbranched laminarin and oligo-laminarins.