Spitting cobras adjust their venom distribution to target distance

If threatened by a human, spitting cobras defend themselves by ejecting their venom toward the face of the antagonist. Circulating head movements of the cobra ensure that the venom is distributed over the face. To assure an optimal distribution of the venom, the amplitudes of head movements should decrease with increasing target distance. To find out whether cobras (Naja pallida and N. nigricollis) adjust their spitting behavior according to target distance we induced spitting from different distances and analyzed their spitting patterns. Our results show that the spray pattern of spiting cobras is not fixed. Instead the snake matches its venom distribution to the size of the target independent of target distance.     

Action mechanism of the platelet aggregation inducer and inhibitor from Echis carinatus snake venom

Platelet aggregation inducer and inhibitor were isolated from Echis carinatus snake venom. The venom inducer caused aggregation of washed rabbit platelets which could be inhibited completely by heparin or hirudin. The venom inducer also inhibit both the reversibility of platelet aggregation induced by ADP and the disaggregating effect of prostaglandin E₁ on the aggregation induced by collagen in the presence of heparin. The venom inhibitor decreased the platelet aggregation induced by collagen, thrombin, ionophore A23187, arachidonate, ADP and platelet-activating factor (PAF) with an IC₅₀ of around 10 μg/ml. It did not inhibit the agglutination of formaldehyde-treated platelets induced by polylysine. In the presence of indomethacin or in ADP-refractory platelets or thrombin-degranulated platelets, the venom inhibitor further inhibited the collagen-induced aggregation. Fibrinogen antagonized competitively the inhibitory action of the venom inhibitor in collagen-induced aggregation. In chymotrypsin-treated platelets, the venom inhibitor abolished the aggregation induced by fibrinogen. It was concluded that the venom inducer caused platelet aggregation indirectly by the conversion of prothrombin to thrombin, while the venom inhibitor inhibited platelet aggregation by interfering with the interaction between fibrinogen and platelets.     

Accelerated Evolution and Molecular Surface of Venom Phospholipase A2 Enzymes

Multiple phospholipase A₂ (PLA₂) isoenzymes found in a single snake venom induce a variety of pharmacological effects. These multiple forms are formed by gene duplication and accelerated evolution of exons. We examined the amino acid sequences of 127 snake venom PLA₂ enzymes and their homologues to study in which location most natural substitutions occur. Our data show that hot spots of amino acid substitutions in this group of proteins occur mostly on the surface. A logistic model correlating the substitution rates of each amino acid residue with their surface accessibility indicates that the probability of natural substitutions occurring in the fully exposed residue is 2.6–3.5 times greater than that of substitutions occurring in buried residues. These surface substitutions play a significant role in the evolution of new PLA₂ isoenzymes by altering the specificity of targeting to various tissues or cells, resulting in distinct pharmacological effects. Thus natural substitutions in PLA₂ enzymes, in contrast to popular belief, are not random substitutions but appear to be directed toward modifying the molecular surface. Received: 11 May 1998 / Accepted: 29 June 1998     

Snake Population Venomics: Proteomics-Based Analyses of Individual Variation Reveals Significant Gene Regulation Effects on Venom Protein Expression in <Emphasis Type="Italic">Sistrurus</Emphasis> Rattlesnakes

Studies of the molecular basis of adaptations seek to understand the relative importance of structural changes in proteins versus gene regulation effects as determinants of phenotype. Amino acid substitutions in gene coding sequences are well documented as causes of variation in snake venom proteins, whereas the importance of gene regulation effects on venom protein abundance and composition is less well known. Here, we use a proteomics-based approach to infer the effects of gene regulation on protein expression by comparing the relative abundance of specific, known venom proteins among different individuals in each of two species of Sistrurus rattlesnakes. Variation in the presence or absence, and in the relative amounts, of proteins was high in both species across all major protein families. Based on our empirical criteria for inferring regulatory effects (presence-absence of specific proteins and/or more than threefold variation in abundance) between 51% and 83% of S. catenatus individuals and between 40% and 63% of S. miliarius individuals showed evidence for gene regulation across the four most abundant proteins (disintegrins, phospholipase A₂’s, serine proteinases, and snake venom metalloproteases). Thus, the effects of gene regulation should be considered an important cause of variation in the composition of whole venoms at the intraspecific level. They also suggest the need for testing the adaptive hypothesis for venom plasticity in relation to prey consumed by adult snakes. Finally, the venom variability reported may have an impact in the treatment of bite victims, highlighting the necessity of using pooled venoms as a substrate for antivenom production.     

Do “infectious” prey select for high levels of natural antibodies in tropical pythons?

Natural antibodies (NAbs) constitute an important component in vertebrate immune system, but, in spite of this, have often been dismissed as “non-specific background” signals. We observed a significant positive relationship between water python (Liasis fuscus) body length/age and levels of antibodies reactive with two administered antigens (tetanus and diphtheria). However, no humoral immune response to the antigens was observed. The lack of elevated immune response, and the age-associated increase in antibody titres, strongly suggest that the antibodies consisted of polyreactive NAbs, and that absence of an elevated immune response was caused by such high levels of NAbs that they were able to mask the epitopes of the antigens. In our study area pythons feed mainly on rodents that frequently, before being killed, are able to inflict numerous bites to the snakes. The bites most likely transmit pathogens such as bacteria. As NAbs have been shown to act as a first line defence against bacterial infections, the high levels of NAbs in the pythons may be an adaptation to reduce pathogenic effects of bacteria transmitted by the prey when the snakes are feeding. Thus, the results from present study suggest that NAbs may have an important immunological function by reducing deleterious effects of pathogens in wild populations.     

Novel Reticular Calcium Binding Protein Is Purified on Taipoxin Columns

Abstract: We identified, by affinity chromatography, two putative binding proteins for the presynaptic snake venom toxin taipoxin. We have previously characterized one of these proteins [neuronal pentraxin (NP)] as a neuronally secreted protein with homology to acute-phase proteins. Here we report the identification of the second protein as a 49-kDa lumenal calcium binding protein that we have named taipoxin-associated calcium binding protein 49 (TCBP-49). This protein contains six EF-hand putative calcium binding domains and the carboxyl-terminal sequence His-Asp-Glu-Leu (HDEL), identical to the yeast endoplasmic reticulum retention signal. Message for this protein is present in brain, liver, muscle, heart, kidney, and testis. Antibodies to this protein label reticular organelles of neurons and glia. This localization and the specific enrichment of native and recombinant TCBP-49 on columns of immobilized taipoxin raise the possibility that this protein interacts with internalized taipoxin, perhaps mediating its activation. The availability of pure TCBP-49 will allow direct tests of whether TCBP-49 alters the integrity of the oligomeric structure, phospholipase activity, or toxicity of taipoxin.     

酶标法观察尖吻蝮出血毒素Ⅰ对毛细血管的作用

从尖吻蝮蛇毒中分离得到的出血毒素Ⅰ(AaHI)具有破坏血管系统并造成广泛出血的作用。本文报道用辣根过氧化物酶标记的AaHI可以特异地结合在毛细血管内皮细胞外膜上,由此推测内皮细胞膜上存在HaHI的受体或作用位点。     

Isolation and characterization of hemorrhagic factors a and b from the venom of the Chinese habu snake (Trimeresurus mucrosquamatus)

Hemorrhagic factors a and b were isolated from Trimeresurus mucrosquamatus venom by Sephadex G-100, CM-Sephadex C-50 and DEAE-Sephacel column chromatographies. The hemorrhagic factors were homogeneous, as established by a single band on acrylamide gel electrophoresis, isoelectric focusing and sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Molecular weights of 15 000 and 27 000 were found for hemorrhagic factors a and b, respectively. Factor a possesses proteolytic activity hydrolyzing the His(10)-Leu(11), Tyr(16)-Leu(17) and Arg(22)-Gly(23) bonds of oxidized insulin B chain, whereas, factor b hydrolyzed only the Ala(14)-Leu(15) bond. Hemorrhagic activity of these hemorrhagic factors was inhibited by ethylenediaminetetraacetic acid, 1,10-phenanthroline or p-chloromercuribenzoate, but not by soybean trypsin inhibitor or diisopropyl fluorophosphate. The hemorrhagic factors were injected into the skin of the back of albino rabbits, and the minimum hemorrhagic dose of factors a and b was 1.7 and 2.3 μg, respectively. These purified hemorrhagic factors were not lethal at 15 μg/g in mice. Factor a hydrolyzed the Bβ chain of fibrinogen, while factor b hydrolyzed the Aα chain. Hemorrhagic factor a was shown to differ immunologically from factor b. Factors a and b produced systemic hemorrhage in internal organs such as the heart and stomach of mice. Moreover, factor b produced hemorrhage in the liver.     

Specific In Vitro Biological Activity of Snake Venom Myotoxins

Abstract: Some snake venoms contain toxins that are reported to be selective for damaging muscle. This specificity can be used to design experiments intended to eliminate muscle. We studied the small myotoxins and fractions IV and V of Bothrops nummifer venom to evaluate their direct effect on cultured muscle cells, neurons, macrophages, and a fibroblast cell line. The small myotoxins, at 100 μg/ml for 2 h, had no effect in vitro, contrary to the in vivo applications. Fractions IV and V were both myotoxic and, at 100 μg/ml, destroyed all cell types. However, at 10 μg/ml the effects of fraction IV were more selective for muscle. Vacuolation of the sarcoplasmic reticulum and T-tubules was first seen in the poisoned muscles, without initial lesions in the nuclei, sarcolemma, mitochondria, and rough endoplasmic reticulum. Fractions IV and V have different toxic activity in cells other than muscles and are a mixture of two basic proteins (i and ii). Protein ii is predominant in fraction IV and protein i is predominant in fraction V. The toxic effects may be mediated by the formation of nonspecific ionic pores in the sarcolemma and/or T-tubule muscle membrane.     

江浙蝮蛇(Agkistrodon halys Pallas)毒的抑菌作用及其抑菌组分L-氨基酸氧化酶的分离纯化

我们用平皿测活法研究了江浙蝮蛇毒对真菌和大肠杆菌生长的抑制作用，并对其抗细菌的组分进行柱层析分离纯化，得到组分单一的LAO不仅有抑菌作用还有L-氨基酸氧化酶的活性。纯化后LAO的比活力为808U／mg。蛇毒经酸性聚丙烯酰胺电泳后测抑菌活性发现有3个抑菌组分(Ⅰ、Ⅱ、Ⅲ)，LAO为抑菌组分Ⅱ。