Glycosylation changes in different developmental stages of Trichinella

The in situ identification of carbohydrate structures in Trichinella spiralis intestinal larvae, adults and L1 muscular larvae was carried out by lectin histochemistry, with emphasis on the O-linked glycans. The absence of reactivity with two lectins-TML and MAL indicated that Trichinella spiralis does not synthesize sialic acid. Reactivity with HPA, VVL-B4, PNA and UEA-I staining suggested that T. spiralis synthesizes and expresses on its cuticle O-linked glycans analogous to Tn-antigen (GalNAc-α-Ser/Thr), T-antigen (Gal-β1,3-GalNAc-α-Ser/Thr) and also structures analogous to A-blood group antigens (GalNAc-α1,3-Gal-β1,3(4)-(Fuc-α1,2-)-R). Expression of the saccharidic moieties is stage-specific. Blood group-A and T-antigen structures were identified on the cuticle of the intestinal and muscular larvae. The Tn-antigen structure was missing in the intestinal larvae. Appropriate ligands for WGA were not identified in the adult individuals. The obtained results may contribute to a better understanding of the glycobiology of this parasitic nematode in relation to occupation of its intracellular niche. The presence of saccharidic structures analogous to some of those expressed on the intestinal epithelial cells may serve as a protective shield on the surface of the parasite.     

Structure of the Human Cation-Independent Mannose 6-Phosphate/IGF2 Receptor Domains 7–11 Uncovers the Mannose 6-Phosphate Binding Site of Domain 9

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Exploring the carbohydrate‐binding ability of Canavalia bonariensis lectin in inflammation models

Lectins are a group of proteins of non‐immune origin recognized for their ability to bind reversibly to carbohydrates. Researchers have been intrigued by oligosaccharides and glycoconjugates for their involvement as mediators of complex cellular events and then many biotechnological applications of lectins are based on glycocode decoding and their activities. Here, we report a structural and biological study of a ConA‐like mannose/glucose‐specific lectin from Canavalia bonariensis seeds, CaBo. More specifically, we evaluate the binding of CaBo with α‐methyl‐D‐mannoside (MMA) and mannose‐1,3‐α‐D‐mannose (M13) and the resultant in vivo effects on a rat model of acute inflammation. A virtual screening was also carried out to cover a larger number of possible bindings of CaBo. In silico analysis demonstrated the stability of CaBo interaction with mannose‐type ligands, and the lectin was able to induce acute inflammation in rats with the participation of the carbohydrate recognition domain (CRD) and histamine release. These results confirm the ability of CaBo to interact with hybrid and high‐mannose N‐glycans, supporting the hypothesis that CaBo's biological activity occurs primarily through its interaction with cell surface glycosylated receptors.     

A novel pattern recognition protein of the Chinese oak silkmoth,Antheraea pernyi,is involved in the pro-PO activating system

In this paper, we firstly reported a C-type lectin cDNA clone of 1029 bps from the larvae of A. Pernyi (Ap-CTL) using PCR and RACE techniques. The full-length cDNA contains an open reading frame encoding 308 amino acid residues which has two different carbohydrate-recognition domains (CRDs) arranged in tandem. To investigate the biological activities in the innate immunity, recombinant Ap-CTL was expressed in E. coli with a 6-histidine at the amino-terminus (Ap-rCTL). Besides acted as a broad-spectrum recognition protein binding to a wide range of PAMPs and microorganisms, Ap-rCTL also had the ability to recognize and trigger the agglutination of bacteria and fungi. In the proPO activation assay, Ap-rCTL specifically restored the PO activity of hemolymph blocked by anti-Ap-rCTL antibody in the presence of different PAMPs or microorganisms. In summary, Ap-rCTL plays an important role in insect innate immunity as an pattern recognition protein. [BMB Reports 2013; 46(7): 358-363]     

Ligand and pathogen specificity of the Atlantic salmon serum C-type lectin

Background

An Atlantic salmon (Salmo salar) C-type lectin (SSL) binds to mannose and related sugars as well as to the surface of Aeromonas salmonicida. To characterize this lectin as a pathogen recognition receptor in salmon, aspects of its interaction with molecules and with intact pathogens were investigated.

Methods

SSL was isolated using whole-yeast-affinity and mannan-affinity chromatography. The binding of SSL to the two major surface molecules of A. salmonicida, lipopolysaccharide (LPS) and A-layer protein was investigated by western blotting and enzyme-linked immunosorbent assays. Microbial binding specificity of SSL was examined by whole cell binding assays using a range of species. Carbohydrate ligand specificity of SSL was examined using glycan array analysis and frontal affinity chromatography.

Results

SSL showed binding to bacteria and yeast including, Pseudomonas fluorescens, A. salmonicida, A. hydrophila, Pichia pastoris, and Saccharomyces cerevisiae, but there was no detectable binding to Yersinia ruckeri. In antimicrobial assays, SSL showed no activity against Escherichia coli, Bacillus subtilis, S. cerevisiae, or A. salmonicida, but it was found to agglutinate E. coli. The major surface molecule of A. salmonicida recognized by SSL was shown to be LPS and not the A-layer protein. LPS binding was mannose-inhibitable. Glycans containing N-acetylglucosamine were shown to be predominant ligands.

Conclusion

SSL has a distinct ligand preference while allowing recognition of a wide variety of related carbohydrate structures.

General Significance

SSL is likely to function as a wide-spectrum pattern recognition protein.     

Expression of integrins α3β1 and α5β1 and GlcNAc β1,6 glycan branching influences metastatic melanoma cell migration on fibronectin

Acquisition of metastatic potential is accompanied by changes in cell surface N-glycosylation. One of the best-studied changes is increased expression of N-acetylglucosaminyltransferase V enzyme (GnT-V) and its products, β1,6-branched N-linked oligosaccharides, observed in the tumorigenesis of many cancers. In this study we demonstrate that during the transition from the vertical growth phase (VGP) (WM793 cell line) to the metastatic stage (WM1205Lu line), β1,6 glycosylation of melanoma cell surface proteins increases as a consequence of elevated expression of the GnT-V-encoding Mgat-5 gene. Treatment with swainsonine led to reduced cell motility on fibronectin in both cell lines; the effect was stronger in metastatic cells, probably due to the higher content of GlcNAc β1,6-branched glycans on the main fibronectin receptors – integrins α5β1 and α3β1. Our results show that GlcNAc β1,6 N-glycosylation of cell surface receptors, which increases with the aggressiveness of melanoma cells, is an important factor influencing melanoma cell migration.     

A rice lectin receptor‐like kinase that is involved in innate immune responses also contributes to seed germination

Seed germination and innate immunity both have significant effects on plant life spans because they control the plant's entry into the ecosystem and provide defenses against various external stresses, respectively. Much ecological evidence has shown that seeds with high vigor are generally more tolerant of various environmental stimuli in the field than those with low vigor. However, there is little genetic evidence linking germination and immunity in plants. Here, we show that the rice lectin receptor‐like kinase OslecRK contributes to both seed germination and plant innate immunity. We demonstrate that knocking down the OslecRK gene depresses the expression of α–amylase genes, reducing seed viability and thereby decreasing the rate of seed germination. Moreover, it also inhibits the expression of defense genes, and so reduces the resistance of rice plants to fungal and bacterial pathogens as well as herbivorous insects. Yeast two‐hybrid and co‐immunoprecipitation experiments revealed that OslecRK interacts with an actin‐depolymerizing factor (ADF) in vivo via its kinase domain. Moreover, the rice adf mutant exhibited a reduced seed germination rate due to the suppression of α–amylase gene expression. This mutant also exhibited depressed immune responses and reduced resistance to biotic stresses. Our results thus provide direct genetic evidence for a common physiological pathway connecting germination and immunity in plants. They also partially explain the common observation that high‐vigor seeds often perform well in the field. The dual effects of OslecRK may be indicative of progressive adaptive evolution in rice.     

Effect of Various Inhibitors on Lipase Action of Thermophilic Fungus Humicola lanuginosa S-38

The hydrolysis of olive oil by the Humicola lipase was inhibited by the addition of n-alcohols, fatty acids and surface active agents. The inhibition of n-alcohols was overcomed by the addition of more substrate but not by the addition of more enzyme. The inhibition of fatty acids and bile salts was eliminated by adding calcium ion. It was concluded that the inhibition of the Humicola lipase by n-alcohols, fatty acids and bile salts was not due to inactivation of the enzyme directly but due to the displacing of the substrate from the oil/water interface, thus blocking the enzyme from the substrate.     

New Metabolites of Streptomyces alboniger

A mutant unable to grow under acidic conditions was isolated from the acidophilic heterotrophic bacterium Acidiphilium facilis 24R. The growth of the mutant could be fully restored by the addition of spermidine or lysine at the concentration of 100 μm. The HPLC analysis of polyamine composition showed that spermidine and putrescine were major polyamine components in the parental strain. In the mutant strain, putrescine was replaced by cadaverine. It was found that some polyamines in the cells were conjugated with the other cell components. The growth of the bacterium in the medium below pH 4.5 was inhibited in the presence of α-methylornithine or methylglyoxal-bis(guanylhydrazone), which are inhibitors of rate-limiting enzymes involved in the biosynthesis of polyamines. The growth of the bacterium that had been inhibited in the presence of inhibitors could be fully restored by the addition of putrescine or spermidine. On the basis of these results, it was concluded that polyamines have a significant role in the growth of Acidiphilium facilis 24R under acidic conditions.