Snake venoms and hemostasis

Snake venoms are complex mixtures of biologically active proteins and peptides. Many of them affect hemostasis by activating or inhibiting coagulant factors or platelets, or by disrupting endothelium. Based on sequence, these snake venom components have been classified into various families, such as serine proteases, metalloproteinases, C-type lectins, disintegrins and phospholipases. The various members of a particular family act selectively on different blood coagulation factors, blood cells or tissues. For almost every factor involved in coagulation or fibrinolysis there is a venom protein that can activate or inactivate it. Venom proteins affect platelet function by binding or degrading vWF or platelet receptors, activating protease-activated receptors or modulating ADP release and thromboxane A2 formation. Some venom enzymes cleave key basement membrane components and directly affect capillary blood vessels to cause hemorrhaging. L-Amino acid oxidases activate platelets via H2O2 production.

Spider Venoms Potentially Lethal to Humans

Spiders have one pair of venom glands, and only a few families have reduced them completely (Uloboridae, Holarchaeidae) or modified them to another function (Symphytognathidae or Scytodidae, see Suter and Stratton 2013). All other 42,000 known spider species (99%) utilize their venom to inject it into prey items, which subsequently become paralysed or are killed. Spider venom is a complex mixture of hundreds of components, many of them interacting with cell membranes or receptors located mainly in the nervous or muscular system (Herzig and King 2013). Spider venom, as it is today, has a 300-million-yearlong history of evolution and adaptation and can be considered as an optimized tool to subdue prey. In Mesothelae, the oldest spider group with less than 100 species, the venom glands lie in the anterior part of the cheliceral basal segment. They are very small and do not support the predation process very effectively. In Mygalomorphae, the venom glands are well developed and fill the basal cheliceral segment more or less completely. Many of these 3,000 species are medium- to large-/very large-sized spiders, and they have created the image of being dangerous beasts, attacking and killing a variety of animals, including humans. Although this picture is completely wrong, it is persistent and contributes considerably to human arachnophobia. The third group of spiders, Araneomorphae or “modern spiders”, comprises 93% of all spider species. The venom glands are enlarged and extend to the prosoma; the openings of the venom ducts are moved from the convex to the concave side of the cheliceral fangs and enlarged as well. These changes save the chelicerae from the necessity of being large, and hence, on the average, araneomorph spiders are much smaller than mygalomorphs. Nevertheless, they possess relatively large venom glands, situated mainly in the prosoma, and may also have rather potent venom.

Main Components of Spider Venoms

Venom glands are alreadypresent in theoldes t spider group, the Mesothelae. Theglands lie in the anterior portion of the cheliceral basal segment but are very small, and it is doubtful how much the venom contributes to the predatory success. In mygalomorph spiders, the well-developed venom glands are still in the basal segment of the chelicerae and produce powerful venom that is injected via the cheliceral fangs into a victim. In all other spiders (Araneomorphae), the venom glands have become much larger and reach into the prosoma where they can take up a considerable proportion of this body part. Only a few spiders have reduced their venom glands, either partially or completely (Uloboridae, Holarchaeidae and Symphytognathidae are usually mentioned) or modified them significantly (Scytodidae, see Suter and Stratton 2013). As well as using venom, spiders may also use their chelicerae to overwhelm an item of prey. It is primarily a question of size whether a spider chews up small arthropods without applying venom or if it injects venom first. Very small and/or defenceless arthropods are picked up and crashed with the chelicerae, while larger, dangerous or well-defended items are carefully approached and only attacked with venom injection. Some spiders specialize on prey groups, such as noctuid moths (several genera of bola spiders among Araneidae), web spiders (Mimetidae), ants (Zodarion species in Zodariidae, aphantochiline thomisids, several genera among Theridiidae, Salticidae, Clubionidae and Gnaphosidae) or termites (Ammoxenidae). However, these more or less monophagous species amount only to roughly 2 % of all known spider species, while 98 % are polyphagous. From these considerations, it follows that the majority of spider venoms are not tailored to any given invertebrate or insect group but are rather unspecialized to be effective over a broad spectrum of prey types that spiders naturally encounter.

The toxins and venoms, and their antibodies,

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Post-translational modification accounts for the presence of varied forms of nerve growth factor in Australian elapid snake venoms

The Australian elapid snakes are amongst the most venomous snakes in the world, but much less is known about the overall venom composition in comparison to Asian and American snakes. We have used a combined approach of cDNA cloning and 2-DE with MS to identify nerve growth factor (NGF) in venoms of the Australian elapid snakes and demonstrate its neurite outgrowth activity While a single 730 nucleotide ORF, coding for a 243 amino acid precursor protein was detected in all snakes, use of 2-DE identified NGF proteins with considerable variation in molecular size within and between the different snakes. The variation in size can be explained at least in part by Winked glycosylation. it is possible that these modifications alter the stability, is necessary to activity and other characteristics of the snake NGFs. Further characterisation delineate the function of the individual NGF isoforms.

BIOCHEMICAL VARIABILITY BETWEEN VENOMS FROM DIFFERENT HONEYBEE (APIS-MELLIFERA) RACES

1. The comparison of molecular exclusion cromatography profiles of venoms from sting apparatuses of Apis mellifera ligustica, Apis mellifera adansonii and Africanized honey-bees in Sephadex G-100 revealed both qualitative and quantitative differences.2. The venoms from A.m. ligustica and A.m. adansonii presented, respectively, three and two peaks characteristic of each sub-species, while Africanized honey-bee was characterized by the absence of eight peaks common to the former.3. The polypeptides with M(r) in the range from 100,000 to 7500 da correspond respectively to 62.0%, 66.6% and 68.7% of total proteins from the venon of A.m. ligustica, A.m. adansonii and Africanized honey-bees, while the peptidic fraction with M(r) range from 4100 to 2000 da corresponds to 11.4%, 32.4% and 10.2% of venom protein, respectively.

The insecticidal spider toxin SFI1 is a knottin peptide that blocks the pore of insect voltage-gated sodium channels via a large β-hairpin loop

Spider venoms contain a plethora of insecticidal peptides that act on neuronal ion channels and receptors. Because of their high specificity, potency and stability, these peptides have attracted much attention as potential environmentally friendly insecticides. Although many insecticidal spider venom peptides have been isolated, the molecular target, mode of action and structure of only a small minority have been explored. Sf1a, a 46-residue peptide isolated from the venom of the tube-web spider Segesteria florentina, is insecticidal to a wide range of insects, but nontoxic to vertebrates. In order to investigate its structure and mode of action, we developed an efficient bacterial expression system for the production of Sf1a. We determined a high-resolution solution structure of Sf1a using multidimensional 3D/4D NMR spectroscopy. This revealed that Sf1a is a knottin peptide with an unusually large β-hairpin loop that accounts for a third of the peptide length. This loop is delimited by a fourth disulfide bond that is not commonly found in knottin peptides. We showed, through mutagenesis, that this large loop is functionally critical for insecticidal activity. Sf1a was further shown to be a selective inhibitor of insect voltage-gated sodium channels, consistent with its 'depressant' paralytic phenotype in insects. However, in contrast to the majority of spider-derived sodium channel toxins that function as gating modifiers via interaction with one or more of the voltage-sensor domains, Sf1a appears to act as a pore blocker.

Lipid composition of the silkworm Bombyx mori L

The constituents of silkworm fat were studied in detail. An unsaturated fat with a high concentration of phospholipid was generally observed. Its iodine value increased during metamorphosis. The free fatty acid concentration likewise increased from the spinning larvae to the moth stage. Analyses of silkworm organs revealed that the fat body had the most fat and the least free fatty acids, whereas haemolymph contained the least fat. Silk glands contained the maximum phospholipid percentage. Stearic acid predominated in those tissues that had a high percentage of phospholipid. Stearic acid was the predominant saturated fatty acid in both the phospholipids and lecithin, and it accounted for 35–50 per cent of the free fatty acids of all the tissues. Q10 was the ubiquinone present; also found were ubichromenol and tocopherol. Results show that silkworm sterol may be cholesterol. Intestines contained the maximum quantities of sterol, ubiquinone, ubichromenol, and tocopherol. The composition of silkworm phospholipids varies considerably from those of other insects, but lecithin is comparable in its composition with lecithins of other animals. The phospholipids had with them a highly complexed protein along with a polysaccharide. In experiments with snake venoms unsaturated fatty acids were found to be predominantly released from silkworm lecithin.

AN ACTIVATOR OF BLOOD-COAGULATION FACTOR-X FROM THE VENOM OF BUNGARUS-FASCIATUS

A specific activator of blood coagulation factor X was purified from the venom of Bungarus fasciatus by gel filtration and by ion-exchange chromatography on a Mono-Q column (FPLC). It consisted of a single polypeptide chain, with a mel. wt of 70,000 in reducing and non-reducing conditions. The enzyme had an amidolytic activity towards the chromogenic substrates S-2266 and S-2302 but it did not hydrolyse S-2238, S2251 or S-2222, which are specific substrates for thrombin, plasmin and factor Xa, respectively. The enzyme activated factor X in vitro and the effect was Ca2+ dependent with a Hill coefficient of 7.9. As with physiological activators, the venom activator cleaves the heavy chain of factor X, producing the activated factor Xa alpha. The purified factor X activator from B. fasciatus venom did not activate prothrombin, nor did it cleave or clot purified fibrinogen. The amidolytic activity and the factor X activation activity of the factor X activator from B. fasciatus venom were readily inhibited by serine protease inhibitors such as diisopropyl fluorophosphate (DFP), phenylmethanesulfonyl fluoride (PMSF), benzamidine and by soybean trypsin inhibitor but not by EDTA. These observations suggest that the factor X activator from B. fasciatus venom is a serine protease. It therefore differs from those of activators obtained from Vipera russelli and Bothrops atrox venoms, which are metalloproteinases.

Effects of Pallas' viper (Agkistrodon halys pallas) venom on blood coagulation and characterization of a prothrombin activator

The action of Pallas' viper (Agkistrodon halys pallas) venom on blood coagulation was examined in vitro and a strong anticoagulant effect was observed. This action was abolished after treatment with a specific inhibitor of phospholipase A(2) activity (p-bromophenacyl bromide), revealing a procoagulant action in low concentrations of treated venom (around 1 mu g/ml). The effect of the venom an haemostasis was further characterized by measuring its ability to activate purified blood coagulation factors. It is concluded that A. halys pallas venom contains prothrombin activation activity. A prothrombin activator (aharin) was purified from the venom by Sephadex G-75 gel filtration and ion-exchange chromatography on a Mono-Q column. It consisted of a single polypeptide chain, with a mol. wt of 63,000. Purified aharin possessed no amidolytic activity on chromogenic substrates. It did not act on other blood coagulation factors, such as factor X and plasminogen, nor did it cleave or clot purified fibrinogen. The prothrombin activation activity of aharin was readily inhibited by ethylenediamine tetracetic acid (a metal chelator), but specific serine protease inhibitors such as diisopropyl fluorophosphate and phenylmethanesulfonyl fluoride had no effect on it. These observations suggest that, like those prothrombin activators from Echis carinatus and Bothrops atrox venoms, the prothrombin activator from A. halys pallas venom is a metalloproteinase. (C) 1998 Elsevier Science Ltd. All rights reserved.

Cloning of cDNAs encoding C-type lectins from Elapidae snakes Bungarus fasciatus and Bungarus multicinctus

A number of C-type lectins with various biological activities have been purified and characterized from Viperidae snake venoms. In contrast, only a few reports could be found in literature concerning the C-type lectins in Elapidae snake venoms. Based on t

The first report of kininogen from invertebrates

The hornet possesses highly toxic venom, which is rich in toxin, enzymes, and biologically active peptides. Several bradykinin-like peptides, vespakinins, have been found in wasp venoms since 1970s, but the mode of biosynthesis of these peptides is unknow