Hemagglutinating Activity Research Articles

Marine bromophenols as an effective inhibitor of virulent proteins (peptidyl arginine deiminase, gingipain R and hemagglutinin A) in Porphyromas gingivalis

ObjectivesPorphyromonas gingivalis, is one of the major oral pathogen that produce virulent proteins which mediate periodontal tissue inflammation and infection. Marine algae have recently gained popularity for its bioactive molecules and their oral applications. Marine bromophenols (MBs) is abundant in red algae which are reported to have wide medicinal properties. The current research primarily focuses to elucidate the bioactivity of MBs against the virulent proteins produced by P. gingivalis. Materials and methodsPotent MBs which effectively binds and inhibits the virulent proteins peptidyl arginine deiminase (PPAD), gingipain R (Rgp) and hemagglutinin A (HgA) was identified through molecular docking and molecular simulation approach. MBs were extracted from Kappaphycus alvarezii (KAB), marine red algae found in India. The efficacy of this MB was studied against P. gingivalis by employing antibacterial activity assays, gingipain assay, hemagglutination inhibition assay (HIA) and mRNA expression analysis (q RT PCR). ResultsMBs with benzene, methyl and glycosyl substitutions demonstrated significant docking score, with good stability and pharmacokinetic properties. In addition to the antibacterial activities against P. gingivalis, KAB was also found to inhibit the gingipain and hemagglutination activities. Exposure of KAB to the virulent genes in P. gingivalis resulted in low mRNA levels of these genes, which suggested the down regulation functions induced by the MBs. ConclusionBiochemical investigations revealed that KAB is a potent natural metabolite that can inhibit and control the virulent proteins produced by P. gingivalis. This study recommends future research to direct towards applicability of MBs in commercial dental products.

Inactivation of lectins from castor cake by alternative chemical compounds

Enabling the use of castor cake in animal feeding is an excellent alternative strategy to reduce feed costs. The cake is a by-product derived from the extraction of the castor oil by the biodiesel industry whose chemical composition is satisfactory despite the presence of antinutritional factors like toxic lectins, which require detoxification before it can be used as a dietary ingredient. The aim of the present study was to evaluate alternative chemical sources in the degradation and inactivation of ricin and Ricinus communis agglutinin (RCA), two lectins from castor cake. Ten chemical compounds were evaluated: sodium hydroxide, monodicalcium phosphate, dicalcium phosphate, calcium oxide, calcium hydroxide, calcitic limestone, magnesian limestone, urea, potassium chloride, and sodium chloride. Gel electrophoresis indicated 100% lectin degradation only in the cakes treated with 90 g sodium hydroxide and 2500 mL water per kg of cake. The hemagglutination assay was crucial to providing innocuousness to the treated cakes, with total absence of hemagglutinating activity observed in the castor cakes treated with 60 or 90 g sodium hydroxide in water volumes equal to or higher than 1500 mL/kg of castor cake and in the cakes treated with 90 g calcium oxide with 2500 or 3000 mL water/kg castor cake. Thus, though depending on the concentration of the chemical compound and on the volume of water per kilogram of treated cake, sodium hydroxide and calcium oxide showed to be promising chemical products for degradation and complete inactivation of the lectins present in castor cake to allow its use as an ingredient in animal diets.

Purification and biophysical characterization of a mannose/N-acetyl-d-glucosamine-specific lectin from Machaerium acutifolium and its effect on inhibition of orofacial pain via TRPV1 receptor

A new mannose/N-acetyl-dglucosamine-specific lectin, named MaL, was purified from seeds of Machaerium acutifolium by precipitation with ammonium sulfate, followed by affinity and ion-exchange chromatography. MaL haemagglutinates either native rabbit erythrocytes or those treated with proteolytic enzymes. MaL is highly stable by the ability to maintain its haemagglutinating activity after exposure to temperatures up to 50 °C. The lectin haemagglutinating activity was optimum between pH 6.0 and 7.0 and inhibited after incubation with d-mannose and N-acetyl-d-glucosamine and α-methyl-d-mannopyranoside. MaL is a glycoprotein with relative molecular mass of 29 kDa (α-chain), 13 kDa (β-chain) and 8 kDa (γ-chain) with secondary structure composed of 3% α-helix, 44% β-sheet, 21% β-turn, and 32% coil. The orofacial antinociceptive activity of the lectin was also evaluated. MaL (0.03 mg mL−1) reduced orofacial nociception induced by capsaicin, an effect that occurred via carbohydrate recognition domain interaction, suggesting an interaction of MaL with the transient receptor potential cation channel subfamily V member 1 (TRPV1) receptor. Our results confirm the potential pharmacological relevance of MaL as an inhibitor of acute orofacial mediated by TRPV1.

The Galactose-Binding Lectin Isolated from Vatairea macrocarpa Seeds Enhances the Effect of Antibiotics Against Staphylococcus aureus-Resistant Strain.

The use of natural products together with standard antimicrobial drugs has recently received more attention as a strategy to combat infectious diseases caused by multidrug-resistant (MDR) microorganisms. This study aimed to evaluate the capacity of a galactose-binding lectin from Vatairea macrocarpa seeds (VML) to modulate antibiotic activity against standard and MDR Staphylococcus aureus and Escherichia coli bacterial strains. The minimum inhibitory concentration (MIC) obtained for VML against all strains was not clinically relevant (MIC ≥ 1024μg/mL). However, when VML was combined with the antibacterial drugs gentamicin, norfloxacin and penicillin, a significant increase in antibiotic activity was observed against S. aureus, whereas the combination of VML and norfloxacin presented decreased and, hence, antagonistic antibiotic activity against E. coli. By its inhibition of hemagglutinating activity, gentamicin (MIC = 50mM) revealed its interaction with the carbohydrate-binding site (CBS) of VML. Using molecular docking, it was found that gentamicin interacts with residues that constitute the CBS of VML with a score of - 120.79 MDS. It is this interaction between the antibiotic and the lectin's CBS that may be responsible for the enhanced activity of gentamicin in S. aureus. Thus, our results suggest that the VML can be an effective modulating agent against S. aureus. This is the first study to report the effect of lectins as modulators of bacterial sensitivity, and as such, the outcome of this study could lay the groundwork for future research involving the use of lectins and conventional antibiotics against such infectious diseases such as community-acquired methicillin-resistant S. aureus (MRSA).

Antitryptical, anticoagulant and hemagglutinating activities of Eucalyptus sp. seeds

Background: Plant biodiversity has great value for science being an inexhaustible source for new bioactive molecules capable of offering environmentally friendly and innovative solutions for various areas of the industry. The scientific community has increased their interest in the study of plant species in the search of new molecules and to determine their mechanisms of action. Plant seeds are natural sources of bioactive compounds, such as carbohydrates, lipids and proteins with special focus on enzymatic inhibitors which protect them against digestive enzymes of phytopathogens and lectins that play an important role on carbohydrate signalization and metabolism during germination. The objective of the present study was to evaluate and describe the protein profile and to test the hemagglutinating, hemolytic and anticoagulant activities, as well as the antitryptic effect of extracts and fractions obtained from seeds of Eucalyptus species. Methods: The crude protein extract was obtained from the seed of Eucalyptus sp. with 0.02 M sodium phosphate buffer, at pH 6.6, and fractionated using ammonium sulfate in order to study its antitryptical properties as well as the capacity of hemagglutination and influence on hemostasis. Results: The crude extract showed a high effectiveness for trypsin inhibition. For hemagglutinating activity, the ammonium sulfate fraction 0-30% presented better activity, while no hemolytic activity was present in the obtained fractions. For anticoagulation assay, the fraction 0-30% showed better results. Conclusions: Taken together, the obtained results demonstrate the biotechnological potential of Eucalyptus sp. seeds, although further study is still necessary to better isolate as well as describe the bioactive compounds.

Three new serine-protease autotransporters of Enterobacteriaceae (SPATEs) from extra-intestinal pathogenic Escherichia coli and combined role of SPATEs for cytotoxicity and colonization of the mouse kidney

ABSTRACTSerine protease autotransporters of Enterobacteriaceae (SPATEs) are secreted proteins that contribute to virulence and function as proteases, toxins, adhesins, and/or immunomodulators. An extra-intestinal pathogenic E. coli (ExPEC) O1:K1 strain, QT598, isolated from a turkey, was shown to contain vat, tsh, and three uncharacterized SPATE-encoding genes. Uncharacterized SPATEs: Sha (Serine-protease hemagglutinin autotransporter), TagB and TagC (tandem autotransporter genes B and C) were tested for activities including hemagglutination, autoaggregation, and cytotoxicity when expressed in E. coli K-12. Sha and TagB conferred autoaggregation and hemagglutination activities. TagB, TagC, and Sha all exhibited cytopathic effects on a bladder epithelial cell line. In QT598, tagB and tagC are tandemly encoded on a genomic island, and were present in 10% of UTI isolates and 4.7% of avian E. coli. Sha is encoded on a virulence plasmid and was present in 1% of UTI isolates and 20% of avian E. coli. To specifically examine the role of SPATEs for infection, the 5 SPATE genes were deleted from strain QT598 and tested for cytotoxicity. Loss of all five SPATEs abrogated the cytopathic effect on bladder epithelial cells, although derivatives producing any of the 5 SPATEs retained cytopathic activity. In mouse infections, sha gene-expression was up-regulated a mean of sixfold in the bladder compared to growth in vitro. Loss of either tagBC or sha did not reduce urinary tract colonization. Deletion of all 5 SPATEs, however, significantly reduced competitive colonization of the kidney supporting a cumulative role of SPATEs for QT598 in the mouse UTI model.

PGN_0297 is an essential component of the type IX secretion system (T9SS) in Porphyromonas gingivalis: Tn-seq analysis for exhaustive identification of T9SS-related genes.

The type IX secretion system (T9SS) was originally discovered in Porphyromonas gingivalis, one of the pathogenic bacteria associated with periodontal disease and is now known to be present in many members of the phylum Bacteroidetes. The T9SS secretes a number of potent virulence factors, including the highly hydrolytic proteases called gingipains, across the outer membrane in P. gingivalis. To understand the entire machinery of T9SS, an exhaustive search for T9SS‐related genes in P. gingivalis using the mariner family transposon (Tn) and Tn‐seq analysis was performed. Seven hundred and two Tn insertion sites in Tn mutants with no colony pigmentation that is associated with Lys‐gingipain (Kgp) defectiveness were determined, and it was found that the Tn was inserted in the kgp gene and 54 T9SS‐related candidate genes. Thirty‐three out of the 54 genes were already known as T9SS‐related genes. Furthermore, deletion mutant analysis of the remaining 21 genes revealed that they were not related to the T9SS. The 33 T9SS‐related genes include a gene for PGN_0297, which was found to be associated with the T9SS components PorK and PorN. A PGN_0297 gene deletion mutant was constructed, and it was found that the mutant showed no colony pigmentation, hemagglutination or gingipain activities, indicating that PGN_0297 was an essential component of the T9SS. The 33 genes did not include the six genes (gppX, omp17, porY, rfa, sigP and wzx) that were also reported as T9SS‐related genes. gppX deletion and insertion mutants were constructed, and it was found that they did not show deficiency in the T9SS.

Portulaca elatior root contains a trehalose-binding lectin with antibacterial and antifungal activities

Lectins are carbohydrate-binding proteins broadly distributed in plants and have several biological functions, including antimicrobial action. Portulaca elatior is a Caatinga plant whose chemical composition and biotechnological potential have not been extensively studied. In this work, a lectin was isolated from P. elatior root extract and evaluated for antimicrobial activity. The P. elatior root lectin (PeRoL) showed native molecular mass of 33 kDa, pI 3.8 and is comprised of two subunits of 15 kDa linked by disulfide bonds. No sequence similarities with Viridiplantae proteins were observed. The PeRoL hemagglutinating activity (HA) was not affected by heating and was detected in a pH ranging from 4.0 to 8.0. Trehalose was identified as an endogenous inhibitor of PeRoL present in the roots. Bacteriostatic activity was detected against Enterococcus faecalis, Pseudomonas aeruginosa and Staphylococcus aureus (minimal inhibitory concentration of 8.1, 32.5 and 4.06 μg/mL, respectively). PeRoL induced the death of Candida albicans, Candida parapsilosis, Candida krusei, and Candida tropicalis cells, with a minimal fungicidal concentration of 16 μg/mL. The lectin (100 μg/mL) was not cytotoxic to human peripheral blood mononuclear cells (PBMCs) and did not show hemolytic activity. In conclusion, the roots of P. elatior contain a trehalose-binding, thermostable, and antimicrobial lectin.

HAEMAGGLUTINATION ACTIVITY OF THE EXTRACTS FROM SOME VIETNAM MARINE INVERTEBRATES

Aqueous extracts from 21 species of Vietnam marine invertebrates, including 11 bivalve and 10 gastropod species, were examined for haemagglutination activity using native and enzyme-treated different animal and human erythrocytes. The 8 bivalve and 10 gastropod species were found to have haemagglutinination activities toward at least one type of erythrocyte tested. A total of 86% of marine invertebrate species surveyed were active. Strong activity was detected in extracts from two bivalve species (Tridacna squamosa and Geloina coaxans) and three gastropod species (Tutufa rubeta, Pleuroploca trapezium and Tectus conus) with enzyme-treated rabbit, horse and human A, B, O erythrocytes. In a haemagglutination–inhibition test with various monosaccharides and glycoproteins, haemagglutination activities of two extracts from T. rubeta and P. trapezium had no affinity for any of the monosaccharides and glycoproteins tested, while activities of the extracts from T. squamosa and T. conus were strongly inhibited by porcine stomach mucin tested, suggesting the presence of lectins specific for O-glycans of these species. The activities of four marine invertebrate extracts were stable over a wide range ofpH and temperature. The haemagglutination activities of T. rubeta and P. trapezium extracts were independent of the presence of divalent cations, whereas the haemagglutination activity of extracts from T. squamosa and T. conus were slightly dependent on the presence of divalent cations. The results suggest that Vietnam marine invertebrates may be good sources of useful lectins for biochemical and biomedical applications.

A new mucin-binding lectin from the marine sponge Aplysina fulva (AFL) exhibits antibiofilm effects

A new mucin-binding lectin (AFL) was isolated from the marine sponge Aplysina fulva. AFL was purified by affinity chromatography on Sepharose™ matrix. Its hemagglutinating activity was independent of divalent ions, and it was weakly inhibited by simple sugars. However, porcine stomach mucin was a powerful inhibitor. In SDS PAGE, piridylethylated AFL showed one band of approximately 16 kDa, whereas in the non-reducing conditions, AFL showed at least two bands of 30 and 70 kDa. Mass spectrometry MALDI-ToF analysis showed one major ion of 31,652 ± 5 Da, which corresponded to a dimer formed by subunits linked by disulfide bonds. The first fifteen amino acids of AFL were determined, and no sequence similarity was observed with any known protein. Internal sequences were obtained by mass spectrometry analysis of tryptic digestion of AFL spots. These peptides showed similarity with a lectin from marine sponge Aplysina lactuca. Secondary structure of AFL was predominantly formed by β-conformations, which were stable at variations of pH and temperature. AFL did not inhibit planktonic growth of Gram-positive and Gram-negative bacteria tested. However, the lectin did significantly reduce the biomass biofilm of the bacteria Staphylococcus aureus, S. epidermidis, and Escherichia coli.

Geodorum densiflorum rhizome lectin inhibits Ehrlich ascites carcinoma cell growth by inducing apoptosis through the regulation of BAX, p53 and NF-κB genes expression

A lectin with a molecular mass of 12 ± 1 kDa was isolated for the first time from Geodorum densiflorum (Lam.) rhizome (GDL). The lectin exhibited hemagglutination activity both in mice and human erythrocytes which was inhibited by 4-nitrophenyl-β-D-glucopyranoside among the tested 26 sugars. The lectin was heat stable and showed its full activity in the pH range from 5.0 to 9.0. The lectin did not lose its activity in the presence of urea but the activity lost significantly when treated with EDTA. Divalent cation Ca2+ and Mg2+ also partially inhibited the activity of the lectin. The lectin strongly agglutinated Ehrlich ascites carcinoma (EAC) and inhibited the cells growth by 60% at 160 μg/ml protein concentration but unable to inhibit the growth of HeLa cells in vitro. The growth inhibition was due to the induction of apoptosis in the EAC cells which was confirmed by annexin-V and caspase-3 substrate and finally by apoptosis-related genes expression. An intensive expression of anti-apoptotic Bcl-X gene was observed only in untreated EAC cells while pro-apoptotic Bak and Bax genes expressed only in GDL treated EAC cells with the remarkable increase of the p53 gene expression. In the treated EAC cells NFκB expression was down regulated.

Isolation and Characterization of a Lectin-like Protein (SBLP) from the Dried Roots of Scutellaria baicalensis (Lamiaceae)

A novel lectin-like protein with MW 63.2 kDa, designated as SBLP, has been isolated and characterized from the dried roots of Scutellaria baicalensis Georgi (Lamiaceae). SBLP was purified by ammonium sulfate precipitation and anion exchange chromatography. It is a glycoprotein according to a PAS staining assay and consisting of protein (86.0%) and sugar (14.0%). Its N-terminal amino acid sequence was determined as GSAVGFLY by Edman degradation. SBLP showed hemagglutinating activity against human and rooster erythrocytes, which were stable below 60°C and in the pH range of 4 −10. Furthermore, SBLP was found to be stimulated by Ca2+, Na+, Ba2+, Zn2+ ions, which suggested it was a metal-dependent lectin. SBLP inhibited the growth of Fusarium oxysporum f.sp. lycopersici and Alternaria eichhorniae in the a dose-dependent manner, and suppressed the proliferation of HepG2 tumor cells with an IC50 of 1.00 μM. This is the first report of a lectin from Radix Scutellariae.

Vigilancia del virus de Influenza Aviar tipo A en patos de crianza familiar de las provincias de Huaral y Huaura (Lima, Perú)

El objetivo del estudio fue evaluar la presencia del virus de influenza aviar (IA) en patos domésticos de crianza familiar en las provincias de Huaral y Huaura (Lima, Perú). Se colectaron 600 muestras de hisopado cloacal de patos domésticos de traspatio, que fueron analizadas mediante aislamiento viral en huevos embrionados de pollo libres de patógenos específicos (SPF). La presencia del virus de IA fue determinada por la actividad hemaglutinante del fluido alantoideo y confirmada mediante un kit comercial que utiliza anticuerpos monoclonales. Todas las muestras analizadas fueron negativas al virus de IA. El riego de infección fue evaluado mediante la simulación de Monte Carlo (programa @risk) determinando que el virus de IA mostraría una prevalencia estocástica máxima de 0.33% con un intervalo de confianza de 95% en caso de presentarse la enfermedad. Se concluye que las aves incluidas en el presente estudio no se encontraban infectadas con el virus de IA.

Induction of Recombinant Lectin Expression by an Artificially Constructed Tandem Repeat Structure: A Case Study Using Bryopsis plumosa Mannose-Binding Lectin.

Lectin is an important protein in medical and pharmacological applications. Impurities in lectin derived from natural sources and the generation of inactive proteins by recombinant technology are major obstacles for the use of lectins. Expressing recombinant lectin with a tandem repeat structure can potentially overcome these problems, but few studies have systematically examined this possibility. This was investigated in the present study using three distinct forms of recombinant mannose-binding lectin from Bryopsis plumosa (BPL2)—i.e., the monomer (rD1BPL2), as well as the dimer (rD2BPL2), and tetramer (rD4BPL2) arranged as tandem repeats. The concentration of the inducer molecule isopropyl β-D-1-thiogalactopyranoside and the induction time had no effect on the efficiency of the expression of each construct. Of the tested constructs, only rD4BPL2 showed hemagglutination activity towards horse erythrocytes; the activity of towards the former was 64 times higher than that of native BPL2. Recombinant and native BPL2 showed differences in carbohydrate specificity; the activity of rD4BPL2 was inhibited by the glycoprotein fetuin, whereas that of native BPL2 was also inhibited by d-mannose. Our results indicate that expression as tandem repeat sequences can increase the efficiency of lectin production on a large scale using a bacterial expression system.

Isolation, Purification and Characterisation of a D-galactose and N-acetyl-D-galactosamine Specific Lectin from Marine Sponge Fasciospongia cavernosa.

Marine sponges, belonging to the phylum Porifera, are gaining more attention by researchers and industrial sectors from all over the world due to their ability to produce a variety of bioactive secondary metabolites that have many applications including drug discovery. Marine sponges are a promising source of bioactive lectins, which are structurally diverse, many of them in the form of glycoproteins. To purify and characterize lectin from a marine sponge Fasciospongia cavernosa and to study its antibacterial activity. Lectin from a marine sponge Fasciospongia cavernosa was purified by guar gum affinity chromatography and checked for its biophysical and antibacterial properties. The lectin was subjected to evaluation for inhibition of microbial growth against bacteria by aggregation test. The activity of FCL against the biofilms formed by P. aeruginosa was also carried out. Biofilm is defined as the undesirable accumulation of microorganisms on artificial surfaces immersed in a common matrix. The effect of FCL on biofilm-forming gram negative bacteria P. aeruginosa was tested by crystal violet assay. This lectin, named FCL, has a molecular weight of 80 KDa approximately, was found to agglutinate human ABO, rat, rabbit and chicken erythrocytes. The hemagglutinating activity of FCL was reduced by demetallisation with E.D.T.A and regained by the addition of Ca2+, Mg2+, Mn2+, Ba2+ and Fe2+, which shows the metal dependency of the lectin. The hemagglutinating activity by the lectin was inhibited by D-galactose and N-acetyl-D-galactosamine. The lectin was stable over a range of pH from 2 to 10.5, and up to a temperature 70°C for 20 min. FCL agglutinated B. subtilis, S. aureus and P. aeruginosa and was able to reduce biofilm mass formed by P. aeruginosa. Thus, the marine sponge F. cavernosa lectin, FCL could be used as an antibacterial agent. FCL significantly reduced the biomass of bacterial biofilm tested. Biofilm mass of P. aeruginosa, K. pneumoniae and E. coli were decreased in a dose dependent manner. A novel lectin was isolated and purified from marine sponge F. cavernosa. FCL, a galactose-binding lectin displayed considerable antimicrobial activity in vitro, particularly against gram-positive bacteria and also exhibited a strong antibiofilm activity.

Isolation and characterization of a galactose-specific lectin (EantH) with antimicrobial activity from Euphorbia antiquorum L. latex

A homodimeric 75 kDa lectin with hemagglutination activity (HA) was purified from the crude latex of Euphorbia antiquorum L. by two types of chromatography, on cation exchange (HiTrap SP FF) and hydrophobic HiTrap Phenyl FF (high sub) columns. The purified protein was designated EantH, and is classified as a galactose-specific thermostable lectin. The HA of EantH was stable at pH values of 5–9 and temperature 5–65 °C. The lectin had bacteriostatic action on the Gram-positive bacteria Staphylococcus aureus and S. epidermidis, with a minimum inhibitory concentration (MIC) of 2000 μg/ml and on a Gram-negative bacterium Samonella typhimurium, with a MIC of 1000 μg/ml. EantH inhibited the growth of Propionibacterium acnes and Streptococcus agalactiae with MIC of 125 μg/ml and 250 μg/ml, respectively. EantH killed P. acnes and S. agalactiae with a minimum microbicidal concentration (MMC) of 1000 μg/ml and 2000 μg/ml, respectively. Scanning electron microscopy indicated that binding of EantH to the carbohydrates in the cell walls of P. acnes and S. typhimurium drastically altered the bacterial cells, and led to inhibition of growth and/or cell death. The antimicrobial activity of EantH could be neutralized by d‑galactose, indicating that its bactericidal action involves binding to galactose in the cell wall.

Identification of key amino acid residues determining ligand binding specificity, homodimerization and cellular distribution of human galectin-10.

Charcot-Leyden crystal protein/Gal-10, abundantly expressed in eosinophils and basophils, is related to several immune diseases. Recently, crystallographic and biochemical studies showed that Gal-10 cannot bind lactose, because a glutamate residue (Glu33) from another monomer blocks the binding site. Moreover, Gal-10 actually forms a novel dimeric structure compared to other galectins. To investigate the role that Glu33 plays in inhibiting lactose binding, we mutated this residue to glutamine, aspartate, and alanine. The structure of E33A shows that Gal-10 can now bind lactose. In the hemagglutination assay, lactose could inhibit E33A from inducing chicken erythrocyte agglutination. Furthermore, we identified a tryptophan residue (Trp127) at the interface of homodimer that is crucial for Gal-10 dimerization. The variant W127A, which exists as a monomer, exhibited higher hemagglutination activity than wild type Gal-10. The solid phase assay also showed that W127A could bind to lactose-modified sepharose-6B, whereas wild type Gal-10 could not. This indicates that the open carbohydrate-binding site of the W127A monomer can bind to lactose. In addition, the distribution of EGFP-tagged Gal-10 and its variants in HeLa cells was investigated. Because Trp72 is the highly conserved in the ligand binding sites of galectins, we used EGFP-tagged W72A to show that Gal-10 could not be transported into the nucleus, indicating that Trp72 is crucial for Gal-10 transport into that organelle.

Hemagglutination ability and hemolytic activity of Trichosporon asahii.

Trichosporon asahii is a human fungal pathogen that causes deep-seated infections in immunocompromised patients. While the pathogenic mechanisms of T. asahii remain unknown, our previous studies indicate that adherent colony morphologies were generated from parent strains, which may contribute to their pathogenicity. In the present study, we analyzed the hemolytic and hemagglutination activities of T. asahii. We report that T. asahii cells demonstrate hemagglutination and hemolytic activities, and that cell surface molecules play a role in the hemagglutination activity of adherent strains. These observations suggest that hemagglutination and hemolysis may be one of the pathogenic mechanisms of T. asahii.

Antiviral Activity of Fermented Ginseng Extracts against a Broad Range of Influenza Viruses.

Ginseng products used as herb nutritional supplements are orally consumed and fermented to ginsenoside compounds by the intestinal microbes. In this study, we investigated antiviral protective effects of fermented ginseng extracts against different strains of influenza viruses in genetically diverse mouse models. Intranasal coinoculation of mice with fermented ginseng extract and influenza virus improved survival rates and conferred protection against H1N1, H3N2, H5N1, and H7N9 strains, with the efficacy dependent on the dose of ginseng samples. Antiviral protection by fermented ginseng extract was observed in different genetic backgrounds of mice and in the deficient conditions of key adaptive immune components (CD4, CD8, B cell, MHCII). The mice that survived primary virus inoculation with fermented ginseng extract developed immunity against the secondary infection with homologous and heterosubtypic viruses. In vitro cell culture experiments showed moderate virus neutralizing activity by fermented ginseng extract, probably by inhibiting hemagglutination and neuraminidase activity. This study suggests that fermented ginseng extracts might provide a means to treat influenza disease regardless of virus strains.

Hericium erinaceus isolectins recognize mucin-type O-glycans as tumor-associated carbohydrate antigens on the surface of K562 human leukemia cells

In Hericium erinaceus mushroom fruiting body, two different lectin groups, HEL1 and HEL2, were identified by using peptide mass fingerprinting based on customized protein sequence databases derived from RNA-Seq data. The HEL2 group included four isoforms designated HEL2a–d. Codon-optimized genes encoding HEL1, HEL2a, and HEL2b were expressed in Escherichia coli to produce fully active soluble proteins designated rHEL1, rHEL2a, and rHEL2b. Interestingly, these lectins showed different molecular weights: approximately 15 kDa for rHEL1 and approximately 120 kDa for rHEL2a and rHEL2b under non-denaturing conditions. rHEL2a and rHEL2b exhibited agglutination activities, but rHEL1 did not show any agglutination activity toward animal erythrocytes. The hemagglutination activity of rHEL2 lectins was strongly inhibited by glycoproteins containing mucin-type O-glycans. Glycan array analysis and isothermal titration calorimetry revealed that rHEL2 isolectins interacted strongly with O-glycans harboring the core 1 O-glycan motif, Galβ(1,3)GalNAc. Moreover, the glycan binding specificities of rHEL2 isolectins were comparable to that of peanut agglutinin in their ability to recognize O-glycans attached to leukosialin as tumor-associated carbohydrate antigens on the surface of K562 human leukemia cells. These results indicate that rHEL2 isolectins could be used as a powerful tool for analyzing mucin-type O-glycans expressed on the surface of cancer cells.