Molecular Characterization and Functional Analysis of the Nattectin-like Toxin from the Venomous Fish Thalassophryne maculosa

TmC4-47.2 is a toxin with myotoxic activity found in the venom of Thalassophryne maculosa, a venomous fish commonly found in Latin America whose envenomation produces an injury characterized by delayed neutrophil migration, production of major pro-inflammatory cytokines, and necrosis at the wound site, as well as a specific systemic immune response. However, there are few studies on the protein structure and functions associated with it. Here, the toxin was identified from the crude venom by chromatography and protein purification systems. TmC4-47.2 shows high homology with the Nattectin from Thalassophryne nattereri venom, with 6 cysteines and QPD domain for binding to galactose. We confirm its hemagglutinating and microbicide abilities independent of carbohydrate binding, supporting its classification as a nattectin-like lectin. After performing the characterization of TmC4-47.2, we verified its ability to induce an increase in the rolling and adherence of leukocytes in cremaster post-capillary venules dependent on the α5β1 integrin. Finally, we could observe the inflammatory activity of TmC4-47.2 through the production of IL-6 and eotaxin in the peritoneal cavity with sustained recruitment of eosinophils and neutrophils up to 24 h. Together, our study characterized a nattectin-like protein from T. maculosa, pointing to its role as a molecule involved in the carbohydrate-independent agglutination response and modulation of eosinophilic and neutrophilic inflammation.

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Molecular Characterization of Rotavirus Porcine OSU Carbohydrate- Binding Domain VP8*

Current Proteomics ◽

10.2174/1570164612666150226232936 ◽

2015 ◽

Vol 12 (1) ◽

pp. 69-72

Author(s):

Hao Li ◽

Wei Zhang ◽

Hong-hao Zhou ◽

Xiao-li Li

Keyword(s):

Molecular Characterization ◽

Binding Domain ◽

Carbohydrate Binding

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Carbohydrate binding activities of Bradyrhizobium japonicum. II. Isolation and characterization of a galactose-specific lectin.

The Journal of Cell Biology ◽

10.1083/jcb.111.4.1639 ◽

1990 ◽

Vol 111 (4) ◽

pp. 1639-1643 ◽

Cited By ~ 25

Author(s):

S C Ho ◽

M Schindler ◽

J L Wang

Keyword(s):

Isoelectric Focusing ◽

Bradyrhizobium Japonicum ◽

Affinity Column ◽

Carbohydrate Binding ◽

Binding Affinities ◽

Isolation And Characterization ◽

Relative Binding ◽

Mannose Derivatives ◽

Derivatives Of

Extracts of Bradyrhizobium japonicum were fractionated on Sepharose columns covalently derivatized with lactose. Elution of the material that was specifically bound to the affinity column with lactose yielded a protein of Mr approximately 38,000. Isoelectric focusing of this sample yielded two spots with pI values of 6.4 and 6.8. This protein specifically bound to galactose-containing glycoconjugates, but did not bind either to glucose or mannose. Derivatives of galactose at the C-2 position showed much weaker binding; there was an 18-fold difference in the relative binding affinities of galactose versus N-acetyl-D-galactosamine. These results indicate that we have purified a newly identified carbohydrate-binding protein from Bradyrhizobium japonicum, that can exquisitely distinguish galactose from its derivatives at the C-2 position.

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Characterization of Oral Squamous Cell Carcinoma Associated Inflammation: A Pilot Study

Frontiers in Oral Health ◽

10.3389/froh.2021.740469 ◽

2021 ◽

Vol 2 ◽

Author(s):

Catherine Laliberté ◽

Nicole Ng ◽

Denise Eymael ◽

Kevin Higgins ◽

Aiman Ali ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Oral Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Immune Cells ◽

Tumor Development ◽

Tissue Analysis ◽

Fluorescent Immunohistochemistry ◽

Pro Inflammatory Cytokines

Background: Oral squamous cell carcinoma (OSCC) is a devastating disease that is usually associated with a dense associated inflammatory infiltrate. Characterizing tumor-associated inflammation is critical to understand the pathogenies of tumor development and progression.Methods: We have tested a protocol to analyze tissue and salivary immune cells and mediators of 37 patients with OSCC at different stages and compared to eight chronic periodontitis patients and 24 healthy controls. Tissue analysis was based on fluorescent immunohistochemistry (FIHC) and inflammatory mediators were analyzed using a Luminex-based 30-Plex panel. Immune cells were analyzed using multichannel flow cytometry including CD45, CD66b, CD3, CD4, CD8, CD25, CD56, CD68, CD138, PD-1, and PD-L1.Results: We show an increase in OSCC-associated inflammation characterized by increased pro-inflammatory cytokines including IL-6, IL-8, TNFα, and GMCSF and increased salivary immune cells.Conclusion: We described a new method to analyze salivary inflammatory markers that can be used in future studies to monitor disease progression and prognosis.

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Affinity purification and affinity characterization of carbohydrate-binding proteins in bovine kidney

Journal of Chromatography A ◽

10.1016/0021-9673(92)80127-g ◽

1992 ◽

Vol 597 (1-2) ◽

pp. 323-330 ◽

Cited By ~ 7

Author(s):

Kyoko Kojima ◽

Haruko K. Ogawa ◽

Nobuko Seno ◽

Isamu Matsumoto

Keyword(s):

Binding Proteins ◽

Affinity Purification ◽

Carbohydrate Binding ◽

Bovine Kidney ◽

Carbohydrate Binding Proteins

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Histomorphological Characterization of Carbohydrate-Binding Proteins in Human Lung Cancer

Lectins and Glycoconjugates in Oncology ◽

10.1007/978-3-642-73662-9_12 ◽

1988 ◽

pp. 131-142 ◽

Cited By ~ 2

Author(s):

K. Kayser ◽

H.-J. Gabius

Keyword(s):

Lung Cancer ◽

Binding Proteins ◽

Human Lung ◽

Human Lung Cancer ◽

Carbohydrate Binding ◽

Carbohydrate Binding Proteins

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Identification and Characterization of a Novel Thermostable and Salt-Tolerant β-1,3 Xylanase from Flammeovirga pacifica Strain WPAGA1

Biomolecules ◽

10.3390/biom10091287 ◽

2020 ◽

Vol 10 (9) ◽

pp. 1287

Author(s):

Zhiwei Yi ◽

Zhengwen Cai ◽

Bo Zeng ◽

Runying Zeng ◽

Guangya Zhang

Keyword(s):

Thermal Stability ◽

Salt Tolerance ◽

3D Structure ◽

Catalytic Efficiency ◽

Dynamics Simulation ◽

Carbohydrate Binding ◽

Salt Tolerant ◽

Excellent Thermal Stability ◽

Moderately Thermophilic

β-1,3 xylanase is an important enzyme in the biorefinery process for some algae. The discovery and characterization of new β-1,3 xylanase is a hot research topic. In this paper, a novel β-1,3 xylanase (Xyl88) is revealed from the annotated genome of Flammeovirga pacifica strain WPAGA1. Bioinformatic analysis shows that Xyl88 belongs to the glycoside hydrolase 26 (GH26) with a suspected CBM (carbohydrate-binding module) sequence. The activity of rXyl88 is 75% of the highest enzyme activity (1.5 mol/L NaCl) in 3 mol/L NaCl buffer, which suggests good salt tolerance of rXy188. The optimum reaction temperature in the buffer without NaCl and with 1.5 mol/L NaCl is 45 °C and 55 °C, respectively. Notably, the catalytic efficiency of rXyl88 (kcat/Km) is approximately 20 higher than that of the thermophilic β-1,3 xylanase that has the highest catalytic efficiency. Xyl88 in this study becomes the most efficient enzyme ever found, and it is also the first reported moderately thermophilic and salt-tolerant β-1,3 xylanase. Results of molecular dynamics simulation further prove the excellent thermal stability of Xyl88. Moreover, according to the predicted 3D structure of the Xyl88, the surface of the enzyme is distributed with more negative charges, which is related to its salt tolerance, and significantly more hydrogen bonds and Van der Waals force between the intramolecular residues, which is related to its thermal stability.

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Chemical studies of Formosan cobra (Naja naja atra) venom. Part I. Chromatographic separation of crude venom on CM-Sephadex and preliminary characterization of its components

Toxicon ◽

10.1016/0041-0101(67)90142-0 ◽

1967 ◽

Vol 5 (1) ◽

pp. 68

Author(s):

S.L.F.

Keyword(s):

Chromatographic Separation ◽

Crude Venom ◽

Naja Naja ◽

Preliminary Characterization ◽

Chemical Studies ◽

Naja Atra

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Cloning and Characterization of Two Xyloglucanases from Paenibacillus sp. Strain KM21

Applied and Environmental Microbiology ◽

10.1128/aem.71.12.7670-7678.2005 ◽

2005 ◽

Vol 71 (12) ◽

pp. 7670-7678 ◽

Cited By ~ 57

Author(s):

Katsuro Yaoi ◽

Tomonori Nakai ◽

Yoshiro Kameda ◽

Ayako Hiyoshi ◽

Yasushi Mitsuishi

Keyword(s):

Glycoside Hydrolase ◽

Amino Acid Sequences ◽

Glycoside Hydrolase Family ◽

Carbohydrate Binding ◽

Carbohydrate Binding Module ◽

C Terminus ◽

Paenibacillus Sp ◽

Genes Encoding ◽

Molecular Masses

ABSTRACT Two xyloglucan-specific endo-β-1,4-glucanases (xyloglucanases [XEGs]), XEG5 and XEG74, with molecular masses of 40 kDa and 105 kDa, respectively, were isolated from the gram-positive bacterium Paenibacillus sp. strain KM21, which degrades tamarind seed xyloglucan. The genes encoding these XEGs were cloned and sequenced. Based on their amino acid sequences, the catalytic domains of XEG5 and XEG74 were classified in the glycoside hydrolase families 5 and 74, respectively. XEG5 is the first xyloglucanase belonging to glycoside hydrolase family 5. XEG5 lacks a carbohydrate-binding module, while XEG74 has an X2 module and a family 3 type carbohydrate-binding module at its C terminus. The two XEGs were expressed in Escherichia coli, and recombinant forms of the enzymes were purified and characterized. Both XEGs had endoglucanase active only toward xyloglucan and not toward Avicel, carboxymethylcellulose, barley β-1,3/1,4-glucan, or xylan. XEG5 is a typical endo-type enzyme that randomly cleaves the xyloglucan main chain, while XEG74 has dual endo- and exo-mode activities or processive endo-mode activity. XEG5 digested the xyloglucan oligosaccharide XXXGXXXG to produce XXXG, whereas XEG74 digestion of XXXGXXXG resulted in XXX, XXXG, and GXXXG, suggesting that this enzyme cleaves the glycosidic bond of unbranched Glc residues. Analyses using various oligosaccharide structures revealed that unique structures of xyloglucan oligosaccharides can be prepared with XEG74.

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Characterization of the Two Neurospora crassa Cellobiose Dehydrogenases and Their Connection to Oxidative Cellulose Degradation

Applied and Environmental Microbiology ◽

10.1128/aem.01503-12 ◽

2012 ◽

Vol 78 (17) ◽

pp. 6161-6171 ◽

Cited By ~ 80

Author(s):

Christoph Sygmund ◽

Daniel Kracher ◽

Stefan Scheiblbrandner ◽

Kawah Zahma ◽

Alfons K. G. Felice ◽

...

Keyword(s):

Neurospora Crassa ◽

Enzyme System ◽

Cellulose Degradation ◽

Cellulose Hydrolysis ◽

Carbohydrate Binding ◽

Ph Optimum ◽

Content Type ◽

Polysaccharide Monooxygenase

ABSTRACTThe genome ofNeurospora crassaencodes two different cellobiose dehydrogenases (CDHs) with a sequence identity of only 53%. So far, only CDH IIA, which is induced during growth on cellulose and features a C-terminal carbohydrate binding module (CBM), was detected in the secretome ofN. crassaand preliminarily characterized. CDH IIB is not significantly upregulated during growth on cellulosic material and lacks a CBM. Since CDH IIB could not be identified in the secretome, both CDHs were recombinantly produced inPichia pastoris. With the cytochrome domain-dependent one-electron acceptor cytochromec, CDH IIA has a narrower and more acidic pH optimum than CDH IIB. Interestingly, the catalytic efficiencies of both CDHs for carbohydrates are rather similar, but CDH IIA exhibits 4- to 5-times-higher apparent catalytic constants (kcatandKmvalues) than CDH IIB for most tested carbohydrates. A third major difference is the 65-mV-lower redox potential of the hemebcofactor in the cytochrome domain of CDH IIA than CDH IIB. To study the interaction with a member of the glycoside hydrolase 61 family, the copper-dependent polysaccharide monooxygenase GH61-3 (NCU02916) fromN. crassawas expressed inP. pastoris. A pH-dependent electron transfer from both CDHs via their cytochrome domains to GH61-3 was observed. The different properties of CDH IIA and CDH IIB and their effect on interactions with GH61-3 are discussed in regard to the proposedin vivofunction of the CDH/GH61 enzyme system in oxidative cellulose hydrolysis.

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