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1.
Purification and amino-acid sequence of a nerve growth factor from the venom of Vipera russelli russelli. 总被引:1,自引:0,他引:1
Nerve growth factor (NGF) was purified from the venom of Vipera russelli russelli by Sephadex G-50 gel filtration, S-Sepharose column chromatography and Blue-Sepharose CL-6B column chromatography. The purified NGF was found to be a glycoprotein, whose apparent molecular mass was estimated to be about 17.5 kDa by SDS-PAGE. The amino-acid sequence was determined by a combination of conventional methods. The V. r. russelli NGF was composed of 117 amino-acid residues with one residue, Asn-21, being N-linked glycosylated and the molecular mass of its protein portion was calculated to be 13,280 Da. 相似文献
2.
Kole L Chakrabarty D Datta K Bhattacharyya D 《Indian journal of biochemistry & biophysics》2000,37(2):114-120
A haemorrhagic toxin (VRR-12) from Vipera russelli russelli (Russell's viper) venom has been purified by ion-exchange chromatography on CM-Sephadex C-50 followed by size-exclusion HPLC to electrophoretically homogeneous state. It is a 12 kDa single polypeptide having 1 mole of Zn+2 ion. This toxin induces intense intestinal haemorrhage and to a lesser extent skeletal muscle haemorrhage in mice. It does not show detectable proteolytic and esterolytic activity with selected substrates under specified conditions, haemolytic and phospholipase activity. When VRR-12, preincubated with bivalent antiserum against Saw-scaled and Russell's viper venom or EDTA was injected, haemorrhagic activity was not reduced, on the other hand preincubation with phenylmethyl sulphonyl fluoride reduced the activity markedly. Biodistribution studies with 125I VRR-12 show that haemorrhagic manifestation by this toxin is not a direct function of the fraction of the totally administered toxin distributed to that tissue. 相似文献
3.
Neuropharmacological studies on the venom of Vipera russelli. 总被引:1,自引:0,他引:1
Neuropharmacological studies have been conducted on the venom of V. russelli on experimental animals. The venom was found to produce alteration in general behaviour pattern, reduction in spontaneous motility, hypothermia, potentiation of pentobarbitone hypnosis, analgesia, reduction in exploratory behaviour pattern, muscle relaxant action, and suppression of aggressive behaviour. The venom caused a significant increase in brain GABA content in mice. The observations are suggestive of a potent CNS-depressant action of V. russelli venom. 相似文献
4.
Vipera russelli venom induces predominantly neurotoxic, myotoxic necrotic and hemorrhagic symptoms in experimental animals and has several hydrolytic enzyme activities. In this study, V. russelli venom is characterized both as a PLA2 and as a toxin. Anti PL-V Ig (antibodies to a toxic phospholipase A2 VRV PL-V of V. russelli venom) nullifies the toxicity of whole V. russelli venom to a great extent. The neurotoxic symptoms vanish completely in the presence of anti PL-V Ig. The cross reacting components of whole V. russelli venom were removed by precipitating them from whole venom by the addition of anti PL-V Ig. The non-cross reacting components present in the supernatant were checked for toxicity. There was a significant reduction in toxicity. The LD50 value of the supernatant had increased from 4.1 mg/kg body weight to 11.7 mg/kg body weight and it showed about 34% of the total venom phospholipase A2 activity. It had edema forming, hemorrhagic and hemolytic activity but failed to induce neurotoxic, anticoagulant and myotoxic effects. 相似文献
5.
A single chain polypeptide, termed beta-RTX, with an apparent Mr = 9600 has been isolated from the venom of Vipera russelli russelli. It was purified by cation exchange chromatography, followed by preparative isoelectric focusing and chromatofocusing. Purity was confirmed by gel filtration, high performance liquid chromatography, gel electrophoresis, and analytical ultracentrifugation. Amino acid analysis revealed the presence of eight half-cystines, one being located at the NH2 terminus, which are linked to form four intramolecular disulfide bridges. Chromatofocusing revealed some microheterogeneity yielding three isoforms with pI values of 9.3, 9.37, and 9.48, respectively. In its native configuration, beta-RTX was not susceptible to tryptic degradation but was readily digested after reduction and alkylation. beta-RTX possesses weak phospholipase A2 activity and competes with the binding of monoamines and opiate ligands to their respective receptors. No binding to histamine, gamma-aminobutyric acid, benzodiazepine, or muscarinic receptors was observed. In vivo, whereas 100 micrograms/kg intravenous beta-RTX seemed to be without apparent effects in the rat, 10 ng/kg beta-RTX injected intracerebroventricularly caused marked sedation, with full recovery within 3 h. 相似文献
6.
All the neurotoxic phospholipases A2 present in whole Vipera russelli venom were precipitated selectively from other non-neurotoxic phospholipases A2 and non-phospholipases A2 fractions using antibodies (anti PL-V Ig) raised against one of the purified neurotoxic phospholipases A2 (VRV PL-V). These neurotoxins were identified and isolated in their homogeneous form by chromatographic and electrophoretic methods. The present report of selective isolation and purification of all the neurotoxic phospholipases A2 of V. russelli venom is first of its kind. 相似文献
7.
Russell's viper (Daboia/Vipera russelli) venom from different regions of India was subjected to chromatographic, electrophoretic, biochemical and immunological analysis. The elution profiles from ion-exchange chromatography and protein banding pattern from SDS-PAGE showed a significant variation in the constituents of venoms. The acidic proteins are found to be predominant in the venoms of eastern and western regions while basic proteins are the major contributors of the northern and southern regional venoms. The major variation of phospholipases A(2) in the venom samples of India may be described as: southern regional venom is rich in basic, toxic PLA(2) while this activity showed a dramatic decrease as one moves towards west, north and eastern regions of India. In addition, the caseinolytic, TAME-hydrolytic, anticoagulant, oedema-inducing and haemorrhagic activities of the venoms have also varied from one region to another. The muscle specimens of mice injected with venoms of different regions showed variable change in the muscle fibre damage and cell morphology. The eastern regional venom is most lethal among all the venoms. The lethal potencies for four regional venoms vary as: eastern>western>southern>northern. The polyclonal antibodies prepared against the venom of southern region showed cross-reaction with the venoms of other regions, but the extent of cross-reaction and diffusion patterns are different. However, the polyclonal antibodies prepared against southern regional venom showed no protection against lethal toxicity of other regional venoms. 相似文献
8.
Blood coagulation factor V (FV) is a multifunctional protein that circulates in human plasma as a precursor molecule which can be activated by thrombin or activated factor X (FXa) in order to express its cofactor activity in prothrombin activation. FV activation is achieved by limited proteolysis after Arg709, Arg1018, and Arg1545 in the FV molecule. The venoms of Daboia russelli and Daboia lebetina contain a serine protease that specifically activates FV by a single cleavage at Arg1545. We have predicted the three-dimensional structure of these enzymes using comparative protein modeling techniques. The plasminogen activator from Agkistrodon acutus, which shows a high degree of homology with the venom FV activators and for which a high-quality crystallographic structure is available, was used as the molecular template. The RVV-V and LVV-V models provide for the first time a detailed and accurate structure of a snake venom FV activator and explain the observed sensitivity or resistance toward a number of serine protease inhibitors. Finally, electrostatic potential calculations show that two positively charged surface patches are present on opposite sides of the active site. We propose that both FV activators achieve their exquisite substrate specificity for the Arg1545 site via interactions between these exosites and FV. 相似文献
9.
Binding of a Glycera convoluta neurotoxin to cholinergic nerve terminal plasma membranes 总被引:4,自引:1,他引:3
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《The Journal of cell biology》1983,97(6):1737-1744
The crude extract of venom glands of the polychaete annelid Glycera convoluta triggers a large Ca2+-dependent acetylcholine release from both frog motor nerve terminals and Torpedo electric organ synaptosomes. This extract was partially purified by Concanavalin A affinity chromatography. The biological activity was correlated in both preparations to a 300,000-dalton band, as shown by gel electrophoresis. This confirmed previous determinations obtained with chromatographic methods. This glycoprotein binds to presynaptic but not postsynaptic plasma membranes isolated from Torpedo electric organ. Pretreatment of intact synaptosomes by pronase abolished both the binding and the venom- induced acetylcholine release without impairing the high K+-induced acetylcholine release. Pretreatment of nerve terminal membranes by Concanavalin A similarly prevented the binding and the biological response. Binding to Torpedo membranes was still observed in the presence of EGTA. An antiserum directed to venom glycoproteins inhibited the neurotoxin so we could directly follow its binding to the presynaptic membrane. Glycera convoluta neurotoxin has to bind to a ectocellularly oriented protein of the presynaptic terminal to induce transmitter release. 相似文献
10.
The venom glands of the annelid Glycera convoluta contain a neurotoxin which triggers ACh release from frog motor terminals and Torpedo synaptosomes. This neurotoxin binds to presynaptic, but not postsynaptic plasma membranes prepared from Torpedo electric organ. The binding site is an ectocellularly oriented protein. The binding does not require Ca. It is inhibited by pretreatment of the membrane by Concanavalin A. The toxin induced ACh release is Ca-dependent and inhibited by D 600. 相似文献
11.
A glycoprotein fraction islated in poor yield (approx. 0.04%) from the venom of Bothrops atrox contained nerve growth factor. The material had biological activity, stability, the property of anomalous adsorption onto surfaces, apparent molecular weight and sub-unit structure that were similar to those for nerve growth factor that had been previously purified from the venom of Vipera russelli. Antiserum raised against nerve growth factor from Vipera russelli showed definite cross-reactivity with either the material from Bothrops atrox or a nerve growth factor-containing fraction from the venom of Ancistrodon piscivorus piscivorus, while antiserum against nerve growth factor from mouse had little effect on any of these preparations. 相似文献
12.
This study describes the characterization of oxylepitoxin-1 (MW 6789), the first postsynaptic neurotoxin isolated from the venom of the Inland taipan (Oxyuranus microlepidotus), which is the most venomous snake in the world. Oxylepitoxin-1, purified using successive steps of size-exclusion and reverse phase-high performance liquid chromatography, produced concentration-dependent (0.3-1.0 microM) inhibition of nerve-mediated (0.1 Hz, 0.2 ms, supramaximal V) twitches of the chick biventer cervicis nerve-muscle preparation. Taipan antivenom (5units/ml) prevented the neurotoxic activity of whole venom (10 microg/ml), but had no significant effect on oxylepitoxin-1 (1 microM). The toxin-induced inhibition of nerve-mediated twitches was significantly reversed upon washing the tissue at 5 min intervals. Oxylepitoxin-1 (30-300 nM) displayed competitive antagonism at the skeletal muscle nicotinic receptor with a pA(2) value of 7.16+/-0.28 (i.e. approximately 10-fold more potent than tubocurarine). The venom had a high level of PLA(2) activity (765+/-73 micromol/min/mg) while oxylepitoxin-1 displayed no PLA(2) activity. Partial N-terminal sequencing of oxylepitoxin-1 shows high sequence identity (i.e. 93%) to postsynaptic toxins isolated from the venom of the closely related coastal taipan (Oxyuranus scutellatus scutellatus). 相似文献
13.
M R Kimball A Sato J S Richardson L S Rosen B W Low 《Biochemical and biophysical research communications》1979,88(3):950-959
Atomic coordinates determined from a 2.5 Å electron density map are given for erabutoxin b, a sea snake venom postsynaptic neurotoxin. The principal structural features, anti-parallel β pleated sheet, β bulge and β bends are described. The erabutoxin b structure is discussed as structural prototype of this class of homologous curare-mimetic neurotoxins from both land and sea snakes. 相似文献
14.
We have investigated the effects of several snake venoms on the Ca2+-dependent K+ channels of human red cells. A heat-resistant component of the venom of the snake Notechis scutatus irreversibly inhibited Ca2+-dependent K+ transport with a Ki value of 0.1-0.2 micrograms/ml. Metabolic changes of the cells modified the maximal effect of the venom. Binding of the venom required extracellular Ca2+ and was quick, but development of full inhibition required additional time. The effects of the venoms from Notechis scutatus and Leiurus quinquestriatus were additive, suggesting that both venoms act through different mechanisms. Venoms of the snakes Vipera russelli russelli and Oxyuranus scutellatus also inhibited Ca2+-dependent K+ transport with the same characteristics as the Notechis scutatus venom. 相似文献
15.
16.
Toshiya Endo Masanao Oya Kyozo Hayashi Tatsuo Miyazawa 《Journal of Protein Chemistry》1989,8(4):575-581
Temperature dependence of proton nuclear magnetic resonance spectra has been followed for cobrotoxin, a postsynaptic neurotoxin fromNaja naja atra venom. Several aromatic amino-acid residues, including the functionally essential Trp-29 located at the tip of the central loop of the molecule, have been found to undergo a thermal structural transition above the global thermal denaturation temperature. It is suggested that a local structure around these residues behaves somehow independently of the rest of the molecule, and that such structural organization may be favorable for a conformational change of a neurotoxin molecule on binding to acetylcholine receptor. 相似文献
17.
Temperature dependence of proton nuclear magnetic resonance spectra has been followed for cobrotoxin, a postsynaptic neurotoxin fromNaja naja atra venom. Several aromatic amino-acid residues, including the functionally essential Trp-29 located at the tip of the central loop of the molecule, have been found to undergo a thermal structural transition above the global thermal denaturation temperature. It is suggested that a local structure around these residues behaves somehow independently of the rest of the molecule, and that such structural organization may be favorable for a conformational change of a neurotoxin molecule on binding to acetylcholine receptor. 相似文献
18.
Two phospholipases A2 (PLA2s), designated as RV-4 and RV-7 were purified from venom of the Taiwan Russell's viper (Vipera russelli formosensis) by gel-filtration and reverse-phase HPLC. Their primary structures were solved by both protein sequencing and cDNA cloning and sequencing. The cDNA synthesized was amplified by the polymerase-chain reaction using a pair of synthetic oligonucleotide primers corresponding to the N- and the C-terminal flanking regions of the enzymes. The deduced amino acid sequences of RV-4 and RV-7 were 92% identical to those of the vipoxin and vipoxin inhibitor, respectively, from the Bulgarian Vipera a. ammodytes. RV-4 itself was neurotoxic, whereas RV-7 had much lower enzymatic activity and was not toxic. The low enzymatic activity of RV-7 may be attributed to five acidic residues at positions 7, 17, 59, 114 and 119, which presumably impair its binding to aggregated lipid substrates. Based on the sequence comparison among all the known group II PLA2s, residues 6, 12, 76-81, and 119-125 were identified as important for the neurotoxicity. RV-4 and RV-7 exist in the crude venom as heterodimers, which were again formed by mixing together the HPLC-purified RV-4 and RV-7. Moreover, RV-7 inhibited the enzymatic activity of RV-4 in vitro but potentiated its lethal potency and neurotoxicity. It is suggested that RV-7 may facilitate the specific binding of RV-4 to its presynaptic binding sites, probably by preventing its non-specific adsorption. 相似文献
19.
Saikia D Bordoloi NK Chattopadhyay P Choklingam S Ghosh SS Mukherjee AK 《Biochimica et biophysica acta》2012,1818(12):3149-3157
An acidic phospholipase A(2) (RVVA-PLA(2)-I) purified from Daboia russelli venom demonstrated dose-dependent catalytic, mitochondrial and erythrocyte membrane damaging activities. RVVA-PLA(2)-I was non-lethal to mice at the tested dose, however, it affected the different organs of mice particularly the liver and cardiac tissues as deduced from the enzymatic activities measured in mice serum after injection of this PLA(2) enzyme. RVVA-PLA(2)-I preferentially hydrolyzed phospholipids (phosphatidylcholine) of erythrocyte membrane compared to the liver mitochondrial membrane. Interestingly, RVVA-PLA(2)-I failed to hydrolyze membrane phospholipids of HT-29 (colon adenocarcinoma) cells, which contain an abundance of phosphatidylcholine in its outer membrane, within 24h of incubation. The gas-chromatographic (GC) analysis of saturated/unsaturated fatty acids' release patterns from intact mitochondrial and erythrocyte membranes after the addition of RVVA-PLA(2)-I showed a distinctly different result. The results are certainly a reflection of differences in the outer membrane phospholipid composition of tested membranes owing to which they are hydrolyzed by the venom PLA(2)s to a different extent. The chemical modification of essential amino acids present in the active site, neutralization study with polyvalent antivenom and heat-inactivation of RVVA-PLA(2)-I suggested the correlation between catalytic and membrane damaging activities of this PLA(2) enzyme. Our study advocates that the presence of a large number of PLA(2)-sensitive phospholipid domains/composition, rather than only the phosphatidylcholine (PC) content of that particular membrane may determine the extent of membrane damage by a particular venom PLA(2) enzyme. 相似文献
20.
勒氏笛鲷微卫星位点的筛选及特征分析 总被引:8,自引:1,他引:7
采用PCR法快速筛选勒氏笛鲷(Lutjanus russelli)基因组文库, 以获得(CA)n微卫星位点。勒氏笛鲷基因组DNA经限制性内切酶HaeⅢ+ DraⅠ双酶切后, 连接T-载体克隆, 构建基因组文库。以通用引物M13+/-与重复序列引物(CA)15对基因组文库进行筛选, 二次筛选后得到121个可能含有微卫星位点的阳性克隆。进行序列测定, 共获得53个CA(n≥7)重复序列, 重复次数主要分布于7~15(80.77%)。在所得微卫星序列中, 重复单元除CA外, 还观察到单碱基、三碱基、四碱基、五碱基重复单元。根据侧翼序列设计48对引物, 通过优化PCR反应条件, 可获得清晰可重复的目的条带。研究旨在为勒氏笛鲷遗传多样性研究及遗传图谱的构建等奠定基础, 为勒氏笛鲷资源的合理开发利用提供参考。 相似文献