Polysaccharides with multiple biological activities are usually considered as one of the major bioactive compounds in Chinese medicines (CMs). At present, the development of drug and functional foods related to polysaccharides have attracted a great deal of attention due to their great potential effects and diverse action mechanisms. However, quality control of polysaccharides is the bottleneck and a challenge due to their complexity and chemical diversity. Actually, the bioactivities of polysaccharides are closely related to their molecular structures. In order to ensure their safety and efficacy, the development of novel approaches based on the molecular structures for the improvement of quality control of polysaccharides is significantly important. Therefore, in this article, the relationship between biological activities and chemical structures, as well as the action mechanisms of polysaccharides from CMs were summarized first. Furthermore, saccharide mapping, a novel strategy for quality control of bioactive polysaccharides from CMs, was introduced and the application and perspectives were also discussed.
Carbohydrate Sequence ; Drugs, Chinese Herbal ; chemistry ; Molecular Sequence Data ; Polysaccharides ; chemistry ; Quality Control

Lipopolysaccharide (LPS) is a major component of the outer membrane of Gram-negative bacteria. Minute amounts of LPS released from infecting pathogens can initiate potent innate immune responses that prime the immune system against further infection. However, when the LPS response is not properly controlled it can lead to fatal septic shock syndrome. The common structural pattern of LPS in diverse bacterial species is recognized by a cascade of LPS receptors and accessory proteins, LPS binding protein (LBP), CD14 and the Toll-like receptor4 (TLR4)-MD-2 complex. The structures of these proteins account for how our immune system differentiates LPS molecules from structurally similar host molecules. They also provide insights useful for discovery of anti-sepsis drugs. In this review, we summarize these structures and describe the structural basis of LPS recognition by LPS receptors and accessory proteins.
Amino Acid Sequence ; Animals ; Binding Sites ; Carbohydrate Sequence ; Humans ; Immunity, Innate ; Lipopolysaccharides/*chemistry/immunology/pharmacology ; Molecular Sequence Data ; Toll-Like Receptor 4/*chemistry/immunology/metabolism

OBJECTIVETo study the chemical constituents of Periploca calophylla.

METHODVarious chromatographic techniques were used to isolate the constituents, and their structures were identified by spectral and chemical methods.

RESULTTwo oligosaccharides were isolated from the chloroform part of P. calophylla and their structures were identified as 4-O-acetyl-beta-cymaropyranosyl (1-->4)-O-beta-D-cymaropyranosyl(1-->4)-O-beta-D-canaropyranosyl (1-->4)-O-beta-D-cymaropyranosy(1-->4)-O-oleandronic acid-delta-lactone(1), and perisaccharide B (2).

CONCLUSIONCompound 1 is a new compound. Compound 2 is reported for the first time from this plant.

Carbohydrate Sequence ; Magnetic Resonance Spectroscopy ; methods ; Molecular Sequence Data ; Molecular Structure ; Oligosaccharides ; analysis ; isolation & purification ; Periploca ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; methods ; Spectroscopy, Fourier Transform Infrared ; methods

As more bioactivities of oligosaccharides have been elucidated, researches on biosynthesis of oligosaccharides have drawn more concerns in Glycobiology. A lot of enzymatic methods for the synthesis of oligosacchrides have been developed employing recombinant E. coli expressed glycosyltranferase or synthase of nucleotide-sugar. This review focuses on the recent progress in the production of oligosaccharides using bacteria especially by genetically engineered bacteria. The key point concering the oligosaccharides biosynthesis in recombinant E. coli, such as enzyme expression, NDP-sugar provision and biosynthesis pathway, was discussed.
Bacteria ; enzymology ; genetics ; Bacterial Proteins ; genetics ; metabolism ; Carbohydrate Sequence ; Escherichia coli ; genetics ; metabolism ; Genetic Engineering ; methods ; trends ; Glycosyltransferases ; genetics ; metabolism ; Industrial Microbiology ; methods ; trends ; Molecular Sequence Data ; Oligosaccharides ; biosynthesis

Tunicamycin, a potent reversible translocase I inhibitor, is produced by several Actinomycetes species. The tunicamycin structure is highly unusual, and contains an 11-carbon dialdose sugar and an α, β-1″,11'-glycosidic linkage. Here we report the identification of a gene cluster essential for tunicamycin biosynthesis by high-throughput heterologous expression (HHE) strategy combined with a bioassay. Introduction of the genes into heterologous non-producing Streptomyces hosts results in production of tunicamycin by these strains, demonstrating the role of the genes for the biosynthesis of tunicamycins. Gene disruption experiments coupled with bioinformatic analysis revealed that the tunicamycin gene cluster is minimally composed of 12 genes (tunA-tunL). Amongst these is a putative radical SAM enzyme (Tun B) with a potentially unique role in biosynthetic carbon-carbon bond formation. Hence, a seven-step novel pathway is proposed for tunicamycin biosynthesis. Moreover, two gene clusters for the potential biosynthesis of tunicamycin-like antibiotics were also identified in Streptomyces clavuligerus ATCC 27064 and Actinosynnema mirums DSM 43827. These data provide clarification of the novel mechanisms for tunicamycin biosynthesis, and for the generation of new-designer tunicamycin analogs with selective/enhanced bioactivity via combinatorial biosynthesis strategies.
Actinobacteria ; enzymology ; genetics ; Base Sequence ; Biological Assay ; Carbohydrate Sequence ; Carbohydrates ; biosynthesis ; genetics ; Cloning, Molecular ; Gene Deletion ; Gene Library ; High-Throughput Screening Assays ; Molecular Sequence Data ; Multigene Family ; Recombinant Proteins ; biosynthesis ; genetics ; Sequence Analysis, DNA ; Streptomyces ; enzymology ; genetics ; Tunicamycin ; biosynthesis ; chemistry ; genetics

Lipooligosacharide (LOS) of Neisseria gonorrhoeae (gonococci, GC) is involved in the interaction of GC with host cells. Deletion of the alpha-oligosaccharide (alpha-OS) moiety of LOS (lgtF mutant) significantly impairs invasion of GC into epithelial cell lines. GC opacity (Opa) proteins, such as OpaI, mediate phagocytosis and stimulate chemiluminescence responses in neutrophils in part through interaction with members of the carcinoembryonic antigen (CEA) family, which includes CEACAM3 (CD66d), a human neutrophil specific receptor for phagocytosis of bacteria. In the present work, we examined the effects of OpaI-expressing lgtF mutant on phagocytosis by HeLa-CEACAM3 cells and chemiluminescence responses in neutrophils. The results showed that lgtF mutant even expressing OpaI completely lost the ability to promote either phagocytosis mediated by CEACAM3 interaction in HeLa cells or chemiluminescence responses in neutrophils. These data indicated that Opa proteins in the lgtF mutant, which might result from the conformational change, cannot be functional.
Antigens, Bacterial ; chemistry ; genetics ; immunology ; metabolism ; Carbohydrate Sequence ; Carcinoembryonic Antigen ; genetics ; immunology ; Gene Expression Regulation ; HeLa Cells ; Host-Pathogen Interactions ; Humans ; Lipopolysaccharides ; chemistry ; immunology ; Luminescent Measurements ; Mutation ; Neisseria gonorrhoeae ; genetics ; metabolism ; pathogenicity ; Neutrophils ; immunology ; microbiology ; Phagocytosis

AIM: To isolate and characterize the anti-complementary polysaccharide from the root of Bupleurum chinense. METHODS: Bioactivity-guided fractionation and purification was used to obtain the anti-complementary polysaccharide from the hot-water extract of the root of Bupleurum chinense. The polysaccharide was characterized by various chemical and spectral analyses. The anti-complementary activities were evaluated by hemolytic assay in vitro. The action targets were identified in the system with individual complement-depleted sera. RESULTS: A homogeneous polysaccharide BC-PS2 was isolated as an anti-complementary agent. It was identified as a branched polysaccharide with an average molecular weight about 2 000 KDa, composed of Glc, Ara, Gal, and Man in the ratio 3.5 : 2.4 : 2.0 : 1.0, respectively, along with a trace of Rha and Xyl, and only 1.11% of protein. The main linkages of the residues of BC-PS2 include terminal, 1, 6-linked, 1, 3-linked and 1, 3, 6-linked Glcp, terminal and 1, 5-linked Araf, terminal, 1, 4-linked, 1, 6-linked and 1, 4, 6-linked Galp, terminal, and, 1, 4-linked and 1, 4, 6-linked Manp. The bioassay experiments revealed that BC-PS2 inhibited complement activation on both the classical and alternative pathways, with CH50 and AP50 of (0.222 ± 0.013) and (0.356 ± 0.032) mg·mL(-1), respectively. Preliminary mechanism studies indicated that BC-PS2 interacted with C1q, C2, and C9 components. CONCLUSION The results demonstrated that BC-PS2 is an anti-complementary polysaccharide, and should be important constituent of the root of Bupleurum chinense for its application in the treatment of diseases associated with the excessive activation of complement system.
Adult ; Bupleurum ; chemistry ; Carbohydrate Sequence ; Complement Activation ; drug effects ; Complement Inactivating Agents ; chemistry ; isolation & purification ; pharmacology ; Hemolysis ; drug effects ; Humans ; Male ; Molecular Weight ; Plant Extracts ; chemistry ; isolation & purification ; pharmacology ; Plant Roots ; chemistry ; Polysaccharides ; chemistry ; isolation & purification ; pharmacology

Plants are known to be efficient hosts for the production of mammalian therapeutic proteins. However, plants produce complex N-glycans bearing β1,2-xylose and core α1,3-fucose residues, which are absent in mammals. The immunogenicity and allergenicity of plant-specific Nglycans is a key concern in mammalian therapy. In this study, we amplified the sequences of 2 plant-specific glycosyltransferases from Nicotiana tabacum L. cv Bright Yellow 2 (BY2), which is a well-established cell line widely used for the expression of therapeutic proteins. The expression of the endogenous xylosyltranferase (XylT) and fucosyltransferase (FucT) was downregulated by using RNA interference (RNAi) strategy. The xylosylated and core fucosylated N-glycans were significantly, but not completely, reduced in the glycoengineered lines. However, these RNAi-treated cell lines were stable and viable and did not exhibit any obvious phenotype. Therefore, this study may provide an effective and promising strategy to produce recombinant glycoproteins in BY2 cells with humanized N-glycoforms to avoid potential immunogenicity.
Amino Acid Sequence ; Blotting, Western ; Carbohydrate Sequence ; Cell Line ; Cloning, Molecular ; DNA, Complementary ; genetics ; Down-Regulation ; Epitopes ; genetics ; immunology ; Fucose ; metabolism ; Fucosyltransferases ; chemistry ; deficiency ; genetics ; immunology ; Glycoproteins ; chemistry ; genetics ; immunology ; Molecular Sequence Data ; Pentosyltransferases ; chemistry ; deficiency ; genetics ; immunology ; Polysaccharides ; chemistry ; immunology ; Protein Engineering ; methods ; RNA Interference ; Species Specificity ; Tobacco ; cytology ; genetics ; Xylose ; metabolism