The Experts below are selected from a list of 9621 Experts worldwide ranked by ideXlab platform

Ayse Nalbantsoy - One of the best experts on this subject based on the ideXlab platform.

  • investigation of vipera anatolica venom Disintegrin via intracellular uptake with radiolabeling study and cell based electrochemical biosensing assay
    Applied Biochemistry and Biotechnology, 2019
    Co-Authors: Ece Eksin, Ayse Nalbantsoy, Bayram Göçmen, Hale Melis Soylu, Fatma Yurt, Arzum Erdem
    Abstract:

    Snake venoms are a natural biological source that has potential therapeutic value with various protein compounds. Disintegrins originally were discovered as a family of proteins from snake venoms composed of cysteine rich low molecular weight polypeptides. Disintegrins exhibit specific binding and higher affinity toward integrin with potential inhibition of function. Trans-membrane receptors of the integrin family may involve in many pathological conditions such as inflammation and tumor progression with important processes related to invasion and migration. Since Disintegrins have the ability to bind to integrins, they could be used for cancer detection and treatment, and in monitoring of therapy in select cancer types. The main purpose of the study is to investigate Disintegrin containing Vipera anatolica (VAT) crude venom potential for radiolabeling and intracellular uptake as well as electrochemical biosensing assay against U87MG human brain glioblastoma cells. For this purpose, VAT crude venom containing U87MG cell-specific Disintegrin was investigated in terms of radiolabeling and intracellular uptake as well as electrochemical biosensing assay in comparison with echistatin (ECT) Disintegrin in cells. The interaction between VAT crude venom and ECT with HEK293 human non-tumorigenic embryonic kidney cells and glioblastoma U87MG cells was electrochemically investigated using pencil graphite electrodes (PGEs). The interaction of the VAT crude venom and ECT with HEK293 and U87MG cells was detected according to the changes in oxidation signals. Then, VAT crude venom and echistatin were labeled with 131I via iodogen method. Intracellular uptakes of radiolabeled molecules were investigated in U87MG cell line. 131I-VAT can be an agent for imaging of glioblastoma cancer. Further work will focus on the production of large quantities of pure VAT Disintegrin with a biotechnological approach to improving imaging agent.

  • investigating the cytotoxic effects of the venom proteome of two species of the viperidae family cerastes cerastes and cryptelytrops purpureomaculatus from various habitats
    Comparative Biochemistry and Physiology C-toxicology & Pharmacology, 2019
    Co-Authors: Cenk Serhan Ozverel, Bayram Göçmen, Maik Damm, Benjaminflorian Hempel, Robert Sroka, Roderich D Sussmuth, Ayse Nalbantsoy
    Abstract:

    Abstract Animal secretions are of great interest in terms of drug development due to their complex protein and peptide composition. Especially, in the field of therapeutic medications such as anti-cancer drugs snake venoms receive attention. In this study, we address two Viperidae species from various habitats with a particular focus on the cytotoxic potential along with the decomplexation of the venom proteome: the horned desert viper (Cerastes cerastes), native to desert regions of North Africa and the mangrove pit viper (Cryptelytrops purpureomaculatus), found in coastal forests of Southeast Asia. Initial cytotoxic screenings of the crude venoms revealed diverse activity, with the highest effect against SHSY5Y human glioblastoma carcinoma cells compared to other cancerous and non-cancerous cell lines. In-depth cytotoxicity studies of SHSY5Y cells with purified venom fractions revealed heterodimeric Disintegrins from C. cerastes venom, which exerted a high cytotoxic activity with IC50 values from 0.11 to 0.58 μM and a Disintegrin-like effect on SHSY5Y morphology was observed due to cell detachment. Furthermore, two polyproline BPP-related peptides, one PLA2 and a peptide-rich fraction were determined for C. purpureomaculatus with moderate IC50 values between 3 and 51 μM. Additionally, the decryption of the venom proteomes by snake venomic mass spectrometry and comparison of the same species from different habitats revealed slight differences in the composition.

  • comprehensive snake venomics of the okinawa habu pit viper protobothrops flavoviridis by complementary mass spectrometry guided approaches
    Molecules, 2018
    Co-Authors: Maik Damm, Ayse Nalbantsoy, Benjaminflorian Hempel, Roderich D Sussmuth
    Abstract:

    The Asian world is home to a multitude of venomous and dangerous snakes, which are used to induce various medical effects in the preparation of traditional snake tinctures and alcoholics, like the Japanese snake wine, named Habushu. The aim of this work was to perform the first quantitative proteomic analysis of the Protobothrops flavoviridis pit viper venom. Accordingly, the venom was analyzed by complimentary bottom-up and top-down mass spectrometry techniques. The mass spectrometry-based snake venomics approach revealed that more than half of the venom is composed of different phospholipases A2 (PLA2). The combination of this approach and an intact mass profiling led to the identification of the three main Habu PLA2s. Furthermore, nearly one-third of the total venom consists of snake venom metalloproteinases and Disintegrins, and several minor represented toxin families were detected: C-type lectin-like proteins (CTL), cysteine-rich secretory proteins (CRISP), snake venom serine proteases (svSP), l-amino acid oxidases (LAAO), phosphodiesterase (PDE) and 5′-nucleotidase. Finally, the venom of P. flavoviridis contains certain bradykinin-potentiating peptides and related peptides, like the svMP inhibitors, pEKW, pEQW, pEEW and pENW. In preliminary MTT cytotoxicity assays, the highest cancerous-cytotoxicity of crude venom was measured against human neuroblastoma SH-SY5Y cells and shows Disintegrin-like effects in some fractions.

  • Comprehensive Snake Venomics of the Okinawa Habu Pit Viper, Protobothrops flavoviridis, by Complementary Mass Spectrometry-Guided Approaches
    MDPI AG, 2018
    Co-Authors: Maik Damm, Ayse Nalbantsoy, Benjaminflorian Hempel, Roderich D Sussmuth
    Abstract:

    The Asian world is home to a multitude of venomous and dangerous snakes, which are used to induce various medical effects in the preparation of traditional snake tinctures and alcoholics, like the Japanese snake wine, named Habushu. The aim of this work was to perform the first quantitative proteomic analysis of the Protobothrops flavoviridis pit viper venom. Accordingly, the venom was analyzed by complimentary bottom-up and top-down mass spectrometry techniques. The mass spectrometry-based snake venomics approach revealed that more than half of the venom is composed of different phospholipases A2 (PLA2). The combination of this approach and an intact mass profiling led to the identification of the three main Habu PLA2s. Furthermore, nearly one-third of the total venom consists of snake venom metalloproteinases and Disintegrins, and several minor represented toxin families were detected: C-type lectin-like proteins (CTL), cysteine-rich secretory proteins (CRISP), snake venom serine proteases (svSP), l-amino acid oxidases (LAAO), phosphodiesterase (PDE) and 5′-nucleotidase. Finally, the venom of P. flavoviridis contains certain bradykinin-potentiating peptides and related peptides, like the svMP inhibitors, pEKW, pEQW, pEEW and pENW. In preliminary MTT cytotoxicity assays, the highest cancerous-cytotoxicity of crude venom was measured against human neuroblastoma SH-SY5Y cells and shows Disintegrin-like effects in some fractions

  • mass spectrometry guided venom profiling and bioactivity screening of the anatolian meadow viper vipera anatolica
    Toxicon, 2015
    Co-Authors: Bayram Göçmen, Paul Heiss, Daniel Petras, Ayse Nalbantsoy
    Abstract:

    This contribution reports on the first characterization of the venom proteome and the bioactivity screening of Vipera anatolica, the Anatolian Meadow Viper. The crude venom as well as an isolated dimeric Disintegrin showed remarkable cytotoxic activity against glioblastoma cells. Due to the rare occurrence and the small size of this species only little amount of venom was available, which was profiled by means of a combination of bottom-up and top-down mass spectrometry. From this analysis we identified snake venom metalloproteases, cysteine-rich secretory protein isoforms, a metalloprotease inhibitor, several type A2 phospholipases, Disintegrins, a snake venom serine protease, a C-type lectin and a Kunitz-type protease inhibitor. Furthermore, we detected several isoforms of above mentioned proteins as well as previously unknown proteins, indicating an extensive complexity of the venom which would have remained undetected with conventional venomic approaches.

Juan J. Calvete - One of the best experts on this subject based on the ideXlab platform.

  • the continuing saga of snake venom Disintegrins
    Toxicon, 2013
    Co-Authors: Juan J. Calvete
    Abstract:

    Disintegrins, a family of polypeptides released in the venoms of viperid snakes (vipers and rattlesnakes) by the proteolytic processing of multidomain metalloproteinases, selectively block the function of β(1) and β(3) integrin receptors. Few of the proteins isolated and characterized from snake venoms have proven to be more structural and functional versatile than the Disintegrins. Not surprisingly, 25 years after their discovery, our knowledge on the evolutionary history and the molecular determinants modulating the integrin inhibitory activity of Disintegrins still remain fragmentary. This paper highlights some seminal contributions, including personal accounts of pioneer authors, related to basic and applied research on Disintegrins. Investigators have evaluated Disintegrin applications in therapies for a number of pathologies in which integrin receptors play relevant roles, particularly myocardial infarction and inappropriate tumor angiogenesis. Completing the continuing story of the Disintegrin family by applying novel research approaches may hold the key to learn how to use deadly toxins as therapeutic agents.

  • evolution of snake venom Disintegrins by positive darwinian selection
    Molecular Biology and Evolution, 2008
    Co-Authors: Paula Juárez, Inaki Comas, Fernando Gonzalezcandelas, Juan J. Calvete
    Abstract:

    PII-Disintegrins, cysteine-rich polypeptides broadly distributed in the venoms of geographically diverse species of vipers and rattlesnakes, antagonize the adhesive functions of beta(1) and beta(3) integrin receptors. PII-Disintegrins evolved in Viperidae by neofunctionalization of Disintegrin-like domains of duplicated PIII-snake venom hemorrhagic metalloproteinase (SVMP) genes recruited into the venom proteome before the radiation of the advanced snakes. Minimization of the gene (loss of introns and coding regions) and the protein structures (successive loss of disulfide bonds) underpins the postduplication divergence of Disintegrins. However, little is known about the underlying genetic mechanisms that have generated the structural and functional diversity among Disintegrins. Phylogenetic inference and maximum likelihood-based codon substitution approaches were used to analyze the evolution of the Disintegrin family. The topology of the phylogenetic tree does not parallel that of the species tree. This incongruence is consistent with that expected for a multigene family undergoing a birth-and-death process in which the appearance and disappearance of loci are being driven by selection. Cysteine and buried residues appear to be under strong purifying selection due to their role in maintaining the active conformation of Disintegrins. Divergence of Disintegrins is strongly influenced by positive Darwinian selection causing accelerated rate of substitution in a substantial proportion of surface-exposed Disintegrin residues. Global and lineage-specific sites evolving under diversifying selection were identified. Several sites are located within the integrin-binding loop and the C-terminal tail, two regions that form a conformational functional epitope. Arginine-glycine-aspartic acid (RGD) was inferred to represent the ancestral integrin-recognition motif, which emerged from the subgroup of PIII-SVMPs bearing the RDECD sequence. The most parsimonious nucleotide substitution model required for the emergence of all known Disintegrin's integrin inhibitory motifs from an ancestral RGD sequence involves a minimum of three mutations. The adaptive advantage of the emergence of motifs targeting beta(1) integrins and the role of positively selected sites located within nonfunctional Disintegrin regions appear to be difficult to rationalize in the context of a predator-prey arms race. Perhaps, this represents a consequence of the neofunctionalization potential of the Disintegrin domain, a feature that may underlie its recruitment into the venom proteome followed by its successful transformation into a toxin.

  • snake venomics of the brazilian pitvipers bothrops cotiara and bothrops fonsecai identification of taxonomy markers
    Journal of Proteomics, 2008
    Co-Authors: Alexandre K Tashima, Antonio C M Camargo, Libia Sanz, Solange M T Serrano, Juan J. Calvete
    Abstract:

    We report the proteomic characterization of venom of the pitvipers Bothrops cotiara and Bothrops fonsecai. Crude venoms were fractionated by reverse-phase HPLC, followed by SDS-PAGE, N-terminal sequencing, MALDI-TOF mass fingerprinting, and CID-MS/MS. Each venom contained around 30 proteins in the range of 7-110 kDa belonging to only 8 (B. cotiara) and 9 (B. fonsecai) families which may target the hemostatic system, albeit distinctly distributed among the two species. B. cotiara and B. fonsecai share medium-sized Disintegrins, Disintegrin-like/cysteine-rich (DC) fragments, snake venom vascular endothelial growth factor, cysteine-rich secretory proteins, serine proteinases, C-type lectins, l-amino acid oxidase, and Zn(2+)-dependent metalloproteinases. In addition, B. fonsecai expresses a high abundance PLA(2) molecule (13,890 Da), whereas PLA(2) molecules were not detected in B. cotiara's venom. This striking finding is in line with previous biochemical analyses showing the absence of phospholipasic activity in the venom of B. cotiara. The potential adaptive significance of the lack of PLA(2) molecules is enigmatic, and alternative explanations are discussed. B. fonsecai is morphologically extremely similar to B. cotiara. Our comparative proteomic analysis shows that compositional differences between their venoms can be employed as a taxonomy signature for unambiguous species identification independently of geographic origin and morphological characteristics.

  • Loss of introns along the evolutionary diversification pathway of snake venom Disintegrins evidenced by sequence analysis of genomic DNA from Macrovipera lebetina transmediterranea and Echis ocellatus
    Journal of Molecular Evolution, 2007
    Co-Authors: Amine Bazaa, Juan J. Calvete, Robert A. Harrison, Paula Juárez, Neziha Marrakchi, Zakaria Bel Lasfer, Mohamed El Ayeb, Libia Sanz
    Abstract:

    Analysis of cDNAs from Macrovipera lebetina transmediterranea (Mlt) and Echis ocellatus (Eo) venom gland libraries encoding Disintegrins argued strongly for a common ancestry of the messengers of short Disintegrins and those for precursors of dimeric Disintegrin chains. We now report the sequence analysis of Disintegrin-coding genes from these two vipers. Genomic DNAs for dimeric Disintegrin subunits Ml_G1 and Ml_G2 (Mlt) and Eo_D3 ( Eo) contain single 1-kb introns exhibiting the 5'-GTAAG (donor)/3'-AG ( acceptor) consensus intron splicing signature. On the other hand, the short RTS-Disintegrins M1_G3 ( Mlt) and Eo_RTS ( Eo) and the short RGD-Disintegrin ocellatusin (Eo) are transcribed from intronless genomic DNA sequences, indicating that the evolutionary pathway leading to the emergence of short Disintegrins involved the removal of all intronic sequences. The insertion position of the intron within Ml_G1, Ml_G2, and Eo_D3 is conserved in the genes for vertebrate ADAM ( A Disintegrin and metalloproteinase) protein Disintegrinlike domains and within the gene for the medium-size snake Disintegrins halystatins 2 and 3. However, a comparative analysis of currently available Disintegrin(-like) genes outlines the view that a minimization of both the gene organization and the protein structure underlies the evolution of the snake venom Disintegrin family.

  • Molecular Cloning of Echis ocellatus Disintegrins Reveals Non-Venom-Secreted Proteins and a Pathway for the Evolution of Ocellatusin
    Journal of Molecular Evolution, 2006
    Co-Authors: Paula Juárez, Robert A. Harrison, Simon C. Wagstaff, Libia Sanz, Juan J. Calvete
    Abstract:

    We report the cloning and sequence analysis of Echis ocellatus cDNAs coding for dimeric Disintegrin subunits and for the short Disintegrin ocellatusin. All the dimeric Disintegrin subunit messengers belong to the short-coding class, indicating that short messengers may be more widely distributed than previously thought. Mass spectrometric analysis of the HPLC-separated venom proteins was performed to characterize the dimeric Disintegrins expressed in the venom proteome. In addition to previously reported EO4 and EO5 heterodimers, a novel dimeric Disintegrin containing RGD- and KGD-bearing subunits was identified. However, a WGD-containing polypeptide encoded by clone Eo1-1 was not detected in the venom, suggesting the occurrence of larger genomic than proteomic diversity, which could represent part of a non-venom-secreted reservoir of Disintegrin that may eventually acquire physiological relevance for the snake upon changes of ecological niches and prey habits. On the other hand, the realization of the existence of two distinct messengers coding for the short Disintegrin ocellatusin reveals key events of the evolutionary emergence of the short Disintegrin ocellatusin from a short-coding dimeric Disintegrin precursor by two nucleotide mutations.

Cezary Marcinkiewicz - One of the best experts on this subject based on the ideXlab platform.

  • pharmacological aspects of vipera xantina palestinae venom
    Toxins, 2011
    Co-Authors: Tatjana Momic, Cezary Marcinkiewicz, Johannes A Eble, Franziska T Arlinghaus, Hadar Arienzakay, Jeoshua Katzhendler, Philip Lazarovici
    Abstract:

    In Israel, Vipera xantina palestinae (V.x.p.) is the most common venomous snake, accounting for several hundred cases of envenomation in humans and domestic animals every year, with a mortality rate of 0.5 to 2%. In this review we will briefly address the research developments relevant to our present understanding of the structure and function of V.x.p. venom with emphasis on venom Disintegrins. Venom proteomics indicated the presence of four families of pharmacologically active compounds: (i) neurotoxins; (ii) hemorrhagins; (iii) angioneurin growth factors; and (iv) different types of integrin inhibitors. Viperistatin, a α1β1selective KTS Disintegrin and VP12, a α2β1 selective C-type lectin were discovered. These snake venom proteins represent promising tools for research and development of novel collagen receptor selective drugs. These discoveries are also relevant for future improvement of antivenom therapy towards V.x.p. envenomation.

  • cdna cloning and functional expression of jerdostatin a novel rts Disintegrin from trimeresurus jerdonii and a specific antagonist of the α1β1 integrin
    Journal of Biological Chemistry, 2005
    Co-Authors: Libia Sanz, Daniel Monleon, Cezary Marcinkiewicz, Bernardo Celda, Paula Juárez, Runqiang Chen, Alicia Perez, Rebeca Hilario, Yuliang Xiong, Enrique Perezpaya
    Abstract:

    Abstract Jerdostatin represents a novel RTS-containing short Disintegrin cloned by reverse transcriptase-PCR from the venom gland mRNA of the Chinese Jerdons pit viper Trimeresurus jerdonii. The jerdostatins precursor cDNA contained a 333-bp open reading frame encoding a signal peptide, a pre-peptide, and a 43-amino acid Disintegrin domain, whose amino acid sequence displayed 80% identity with that of the KTS-Disintegrins obtustatin and viperistatin. The jerdostatin cDNA structure represents the first complete open reading frame of a short Disintegrin and points to the emergence of jerdostatin from a short-coding gene. The different residues between jerdostatin and obtustatin/viperistatin are segregated within the integrin-recognition loop and the C-terminal tail. Native jerdostatin (r-jerdostatin-R21) and a R21K mutant (r-jerdostatin-K21) were produced in Escherichia coli. In each case, two conformers were isolated. One-dimensional 1H NMR showed that conformers 1 and 2 of r-jerdostatin-R21 represent, respectively, well folded and unfolded proteins. The two conformers of the wild-type and the R21K mutant inhibited the adhesion of α1-K562 cells to collagen IV with IC50 values of 180 and 703 nm, respectively. The IC50 values of conformers 2 of r-jerdostatin-R21 and r-jerdostatin-K21 were, respectively, 5.95 and 12.5 μm. Neither r-jerdostatin-R21 nor r-jerdostatin-K21 showed inhibitory activity toward other integrins, including αIIbβ3, αvβ3, α2β1, α5β1, α4β1, α6β1, and α9β1 up to a concentration of 24 μm. Although the RTS motif appears to be more potent than KTS inhibiting the α1β1 integrin, r-jerdostatin-R21 is less active than the KTS-Disintegrins, strongly suggesting that substitutions outside the integrin-binding motif and/or C-terminal proteolytic processing are responsible for the decreased inhibitory activity.

  • snake venom Disintegrins novel dimeric Disintegrins and structural diversification by disulphide bond engineering
    Biochemical Journal, 2003
    Co-Authors: Juan J. Calvete, Paz M Morenomurciano, David R G Theakston, Dariusz G Kisiel, Cezary Marcinkiewicz
    Abstract:

    We report the isolation and amino acid sequences of six novel dimeric Disintegrins from the venoms of Vipera lebetina obtusa (VLO), V. berus (VB), V. ammodytes (VA), Echis ocellatus (EO) and Echis multisquamatus (EMS). Disintegrins VLO4, VB7, VA6 and EO4 displayed the RGD motif and inhibited the adhesion of K562 cells, expressing the integrin alpha5beta1 to immobilized fibronectin. A second group of dimeric Disintegrins (VLO5 and EO5) had MLD and VGD motifs in their subunits and blocked the adhesion of the alpha4beta1 integrin to vascular cell adhesion molecule 1 with high selectivity. On the other hand, Disintegrin EMS11 inhibited both alpha5beta1 and alpha4beta1 integrins with almost the same degree of specificity. Comparison of the amino acid sequences of the dimeric Disintegrins with those of other Disintegrins by multiple-sequence alignment and phylogenetic analysis, in conjunction with current biochemical and genetic data, supports the view that the different Disintegrin subfamilies evolved from a common ADAM (a Disintegrin and metalloproteinase-like) scaffold and that structural diversification occurred through disulphide bond engineering.

  • amino acid sequence and homology modeling of obtustatin a novel non rgd containing short Disintegrin isolated from the venom of vipera lebetina obtusa
    Protein Science, 2003
    Co-Authors: Paz M Morenomurciano, Daniel Monleon, Cezary Marcinkiewicz, Bernardo Celda, Juan J. Calvete
    Abstract:

    Disintegrins represent a group of cysteine-rich peptides occurring in Crotalidae and Viperidae snake venoms, and are potent antagonists of several integrin receptors. A novel Disintegrin, obtustatin, was isolated from the venom of the Vipera lebetina obtusa viper, and represents the first potent and selective inhibitor of the binding of integrin α1β1 to collagen IV. The primary structure of obtustatin contains 41 amino acids and is the shortest Disintegrin described to date. Obtustatin shares the pattern of cysteines of other short Disintegrins. However, in contrast to known short Disintegrins, the integrin-binding loop of obtustatin is two residues shorter and does not express the classical RGD sequence. Using synthetic peptides, a KTS motif was identified as the integrin-binding sequence. A three-dimensional model of obtustatin, built by homology-modeling structure calculations using different templates and alignments, strongly indicates that the novel KTS motif may reside at the tip of a flexible loop.

  • characterization of a monomeric Disintegrin ocellatusin present in the venom of the nigerian carpet viper echis ocellatus
    FEBS Letters, 2002
    Co-Authors: Bryan J Smith, Juan J. Calvete, David R G Theakston, Ana Lucia Coelho, Christina Barjafidalgo, Cezary Marcinkiewicz
    Abstract:

    Ocellatusin is a new RGD-containing short monomeric Disintegrin. It is a better inhibitor of alpha(5)beta(1) integrin and a more potent inducer of the expression of a ligand-induced binding site epitope on beta(1) integrin subunit than echistatin. In further contrast to echistatin, ocellatusin has a direct chemotactic stimulus on human neutrophils in vitro. The distinct effects of these two close evolutionarily related Disintegrins might be explained by the presence of methionine-22 and histidine-29 in the RGD loop of ocellatusin, which are arginine and aspartic acid, respectively, in echistatin. These mutations may modulate the conformation and/or recognition properties of the integrin-binding loop of ocellatusin.

Roderich D Sussmuth - One of the best experts on this subject based on the ideXlab platform.

  • investigating the cytotoxic effects of the venom proteome of two species of the viperidae family cerastes cerastes and cryptelytrops purpureomaculatus from various habitats
    Comparative Biochemistry and Physiology C-toxicology & Pharmacology, 2019
    Co-Authors: Cenk Serhan Ozverel, Bayram Göçmen, Maik Damm, Benjaminflorian Hempel, Robert Sroka, Roderich D Sussmuth, Ayse Nalbantsoy
    Abstract:

    Abstract Animal secretions are of great interest in terms of drug development due to their complex protein and peptide composition. Especially, in the field of therapeutic medications such as anti-cancer drugs snake venoms receive attention. In this study, we address two Viperidae species from various habitats with a particular focus on the cytotoxic potential along with the decomplexation of the venom proteome: the horned desert viper (Cerastes cerastes), native to desert regions of North Africa and the mangrove pit viper (Cryptelytrops purpureomaculatus), found in coastal forests of Southeast Asia. Initial cytotoxic screenings of the crude venoms revealed diverse activity, with the highest effect against SHSY5Y human glioblastoma carcinoma cells compared to other cancerous and non-cancerous cell lines. In-depth cytotoxicity studies of SHSY5Y cells with purified venom fractions revealed heterodimeric Disintegrins from C. cerastes venom, which exerted a high cytotoxic activity with IC50 values from 0.11 to 0.58 μM and a Disintegrin-like effect on SHSY5Y morphology was observed due to cell detachment. Furthermore, two polyproline BPP-related peptides, one PLA2 and a peptide-rich fraction were determined for C. purpureomaculatus with moderate IC50 values between 3 and 51 μM. Additionally, the decryption of the venom proteomes by snake venomic mass spectrometry and comparison of the same species from different habitats revealed slight differences in the composition.

  • comprehensive snake venomics of the okinawa habu pit viper protobothrops flavoviridis by complementary mass spectrometry guided approaches
    Molecules, 2018
    Co-Authors: Maik Damm, Ayse Nalbantsoy, Benjaminflorian Hempel, Roderich D Sussmuth
    Abstract:

    The Asian world is home to a multitude of venomous and dangerous snakes, which are used to induce various medical effects in the preparation of traditional snake tinctures and alcoholics, like the Japanese snake wine, named Habushu. The aim of this work was to perform the first quantitative proteomic analysis of the Protobothrops flavoviridis pit viper venom. Accordingly, the venom was analyzed by complimentary bottom-up and top-down mass spectrometry techniques. The mass spectrometry-based snake venomics approach revealed that more than half of the venom is composed of different phospholipases A2 (PLA2). The combination of this approach and an intact mass profiling led to the identification of the three main Habu PLA2s. Furthermore, nearly one-third of the total venom consists of snake venom metalloproteinases and Disintegrins, and several minor represented toxin families were detected: C-type lectin-like proteins (CTL), cysteine-rich secretory proteins (CRISP), snake venom serine proteases (svSP), l-amino acid oxidases (LAAO), phosphodiesterase (PDE) and 5′-nucleotidase. Finally, the venom of P. flavoviridis contains certain bradykinin-potentiating peptides and related peptides, like the svMP inhibitors, pEKW, pEQW, pEEW and pENW. In preliminary MTT cytotoxicity assays, the highest cancerous-cytotoxicity of crude venom was measured against human neuroblastoma SH-SY5Y cells and shows Disintegrin-like effects in some fractions.

  • Comprehensive Snake Venomics of the Okinawa Habu Pit Viper, Protobothrops flavoviridis, by Complementary Mass Spectrometry-Guided Approaches
    MDPI AG, 2018
    Co-Authors: Maik Damm, Ayse Nalbantsoy, Benjaminflorian Hempel, Roderich D Sussmuth
    Abstract:

    The Asian world is home to a multitude of venomous and dangerous snakes, which are used to induce various medical effects in the preparation of traditional snake tinctures and alcoholics, like the Japanese snake wine, named Habushu. The aim of this work was to perform the first quantitative proteomic analysis of the Protobothrops flavoviridis pit viper venom. Accordingly, the venom was analyzed by complimentary bottom-up and top-down mass spectrometry techniques. The mass spectrometry-based snake venomics approach revealed that more than half of the venom is composed of different phospholipases A2 (PLA2). The combination of this approach and an intact mass profiling led to the identification of the three main Habu PLA2s. Furthermore, nearly one-third of the total venom consists of snake venom metalloproteinases and Disintegrins, and several minor represented toxin families were detected: C-type lectin-like proteins (CTL), cysteine-rich secretory proteins (CRISP), snake venom serine proteases (svSP), l-amino acid oxidases (LAAO), phosphodiesterase (PDE) and 5′-nucleotidase. Finally, the venom of P. flavoviridis contains certain bradykinin-potentiating peptides and related peptides, like the svMP inhibitors, pEKW, pEQW, pEEW and pENW. In preliminary MTT cytotoxicity assays, the highest cancerous-cytotoxicity of crude venom was measured against human neuroblastoma SH-SY5Y cells and shows Disintegrin-like effects in some fractions

Janice P. Evans - One of the best experts on this subject based on the ideXlab platform.

  • analysis of fertilin α adam1 mediated sperm egg cell adhesion during fertilization and identification of an adhesion mediating sequence in the Disintegrin like domain
    Journal of Biological Chemistry, 2001
    Co-Authors: Grace E Wong, Xiaoling Zhu, Chrissy E Prater, Janice P. Evans
    Abstract:

    Abstract Fertilin α (also known as ADAM1) is a member of the ADAM (A Disintegrin and A metalloprotease domain) family of proteins. In this study, we examine the mechanism of mouse fertilin α's in adhesion of sperm to the egg plasma membrane during fertilization. We find that recombinant forms of fertilin α corresponding to either the Disintegrin-like domain or the cysteine-rich domain and the EGF-like repeat can perturb sperm-egg binding, suggesting that both of these domains can participate in fertilin α-mediated adhesion events. In further examination of the fertilin α Disintegrin-like domain, we find that a subdomain of Disintegrin-like domain with the sequence DLEECDCG outside the putative Disintegrin loop but with homology to the fertilin β Disintegrin loop can inhibit the binding of both sperm and recombinant fertilin α to eggs, suggesting that this is an adhesion-mediating motif of the fertilin α Disintegrin-like domain. This sequence also inhibits the binding of recombinant fertilin β to eggs and thus is the first peptide sequence found to block two different sperm ligands. Finally, a monoclonal antibody to the tetraspanin protein CD9, KMC.8, inhibited the binding of recombinant fertilin α to eggs in one type of binding assay, suggesting that, under certain conditions, fertilin α may interact with a KMC.8-sensitive binding site on the egg plasma membrane.

  • identification of key functional amino acids of the mouse fertilin β adam2 Disintegrin loop for cell cell adhesion during fertilization
    Journal of Biological Chemistry, 2000
    Co-Authors: Xiaoling Zhu, Niharika P Bansal, Janice P. Evans
    Abstract:

    Abstract Fertilin β (also known as ADAM2) is a cell adhesion molecule on the surface of mammalian sperm that participates in sperm-egg membrane binding. Fertilin β is a member of the molecular family known as ADAMs or MDCs. These proteins have a Disintegrin domain with homology to integrin ligands found in snake venoms; several of these snake proteins have an RGD tripeptide presented on an extended “Disintegrin loop.” However, fertilin β lacks an RGD tripeptide and instead has the consensus sequenceX(D/E)ECD (QDECD in mouse fertilin β) in its putative Disintegrin loop, and there is controversy over which amino acids comprise the active site of the fertilin β Disintegrin loop. We have used point-mutated versions of the sequence AQDECDVT and two bioassays to identify the key functional amino acids of this sequence from the mouse fertilin β Disintegrin domain. Amino acid substitutions for the terminal aspartic acid residue of the QDECD sequence result in dramatically reduced activities in the two assays for protein function, implicating the terminal aspartic acid residue as critical for protein function. Substitutions for the glutamic acid and the cysteine residues in the QDECD sequence result in slight reductions in activity, whereas substitution of the first aspartic acid has virtually no effect. These data suggest that the conserved ECD sequence of the mouse fertilin β Disintegrin loop, especially the terminal D residue, contributes more to the protein's activity than does the QDE sequence that aligns with the RGD tripeptide in other Disintegrins.

  • Roles of the Disintegrin domains of mouse fertilins alpha and beta in fertilization
    1998
    Co-Authors: Janice P. Evans, Richard M. Schultz, Gregory S. Kopf
    Abstract:

    Fertilin is a heterodimer of a and b subunits, both of which are members of the ADAM (A Disintegrin and A Metalloprotease do-main)/MDC (Metalloprotease-Disintegrin-Cysteine-rich) family of proteins. We have previously demonstrated that recombinant forms of the putative extracellular domains of mouse fertilin a and fertilin b bind to mouse eggs and inhibit sperm-egg membrane binding. In this study, we examined the roles of the Disintegrin domains of fertilins a and b by producing recombinant forms of fertilins a and b that included the Disintegrin domains (aDCE and bDCE) or that were truncated so that they lack the Disintegrin domains (aCE and bCE) and tested the abilities of these proteins to bind to eggs and to inhibit sperm-egg binding. Fertilin bDCE was able to inhibit sperm-egg binding, but fertilin bCE was relatively ineffective, in-dicating that the Disintegrin domain of fertilin b is required for interactions with egg binding sites and/or for proper protein fold-ing. Fertilins aDCE and aCE both inhibited sperm-egg interactions, but fertilin aDCE tended to be more effective. Thus, the presence of the Disintegrin domain in fertilin aDCE apparently enhanced the ability of this recombinant protein to inhibit sperm-egg binding, either by interacting with egg binding sites or by improving the efficiency of protein folding. These data also indicate that the other domains of the fertilin a extracellular region (cysteine-rich and/or epidermal growth factor-like repeat) have the ability to block sperm binding and suggest that these domains of fertilin a may participate in sperm-egg adhesion

  • mouse sperm egg plasma membrane interactions analysis of roles of egg integrins and the mouse sperm homologue of ph 30 fertilin beta
    Journal of Cell Science, 1995
    Co-Authors: Janice P. Evans, Richard M. Schultz, Gregory S. Kopf
    Abstract:

    The guinea pig sperm protein, PH-30 (also known as fertilin), is postulated to participate in the interaction between the sperm and egg plasma membranes. The beta subunit of guinea pig PH-30 (gpPH-30 beta) contains a domain with homology to Disintegrins, snake venom proteins that bind to integrins via an integrin-binding domain containing the tripeptide RGD. This raises the question of whether an egg integrin serves as a receptor for PH-30. Although mouse eggs express integrin subunits, their role in mouse fertilization is unresolved. Therefore, we examined fertilization for two different hallmarks of integrin function, namely, dependence of ligand binding on divalent cations and the ability to inhibit ligand binding with RGD peptides. We demonstrate that sperm binding to zona pellucida-free eggs is supported by Ca2+, Mg2+, or Mn2+. Ca2+ was necessary and sufficient for sperm-egg fusion, with 2.5 mM Ca2+ being the most effective concentration. In addition, fertilization could be partially inhibited with various RGD peptides, which caused a decrease in sperm-egg fusion by 30–58%. This partial inhibition of fusion with RGD peptides prompted the cloning of the mouse homologue of gpPH-30 beta (hereafter referred to as mPH-30 beta) to determine if it possessed the tripeptide RGD or a different amino acid sequence in its Disintegrin domain. mPH-30 beta, which is expressed during meiotic and post-meiotic phases of spermatogenesis, shares significant similarities to gpPH-30 beta throughout the length of the molecule, from the signal sequence to the cytoplasmic tail. The full-length deduced amino acid sequence of mPH-30 beta. The Disintegrin domain of mPH-30 beta has the tripeptide QDE (instead of RGD) in its cell recognition region. Peptides containing this QDE sequence decrease the binding and fusion of sperm with zona pellucida-free eggs by approximately 70%, suggesting that the Disintegrin domain of mPH-30 beta participates in the interaction between sperm and egg membranes.