Pharmacological and immunological evaluation of scorpion (Heterometrus bengalensis) venom antiserum

An antiserum was prepared for the first time against the venom of a common scorpion, H. bengalensis, by hyperimmunization of rabbit. This antiserum showed positive precipitin bands in immunogeldiffusion and immunoelectrophoresis. The serum showed a high titre value tested by indirect haemagglutination test. The antiserum developed in rabbit protected mice against the lethal action of the venom. Smooth muscle contractile response of venom on guinea pig ileum, and rat uterus was antagonized by the antiserum. This antiserum effectively antagonized the venom induced neuromuscular paralysis tested on rat phrenic nerve diaphragm and chick biventer cervices. Antiserum also protected the venom-induced cardiac arrest tested on isolated guineapig heart and auricle preparations.     

Isolation and characterization of hyaluronidase from scorpion (Heterometrus fulvipes) venom

Hyaluronidase (Hyaluronate lyase, E.C. 3.2.1.35) has been isolated from Heterometrus fulvipes scorpion venom by a combination of gel filtration on Sephadex G-75 and ion exchange chromatography on DEAE-cellulose. The enzyme preparation showed a single band on polyacrylamide gel electrophoresis and a molecular weight of 82,000. The final preparation was purified 27-fold. The optimum pH for enzyme activity was 4.0. No loss of activity was observed up to 30 degrees C and showed a sharp decrease in activity at 50 degrees C. Heparin inhibited the enzyme activity.     

Comparative detoxification and protection of scorpion (Heterometrus bengalensis) venom by toxoid antiserum

Comparative detoxification of scorpion venom by using different chemical agents was investigated. Detoxification by formalin showed the best optimum detoxifying agent. The formalin treatment resulted in 2.3% protein loss with 6-fold detoxification. This formal toxoid was immunogenic in rabbit giving high neutralizing antibodies as revealed from indirect haemagglutination test. Toxoid antiserum protected mice against the lethal action of venom. It also effectively antagonized the smooth muscle contractile response of venom, and venom-induced neuromuscular paralysis. This toxoid antiserum also protected the venom-induced cardiac arrest. The possibility of using this formal toxoid for antisera production and immunization for therapeutic use needs to be explored.     

Detection and distribution of scorpion (Heterometrus bengalensis) venom in rabbit tissues by enzyme-linked immunosorbent assay (ELISA).

ELISA was carried out to detect distribution of scorpion venom in experimental animal tissues. The venom content of different tissues was in the order, liver greater than kidney greater than spleen greater than lung greater than heart greater than diaphragm greater than brain. Tissue distribution of venom antigen in the envenomental subject by ELISA will provide a better approach for serotherapy.     

Rhythmic changes in the activity of neurosecretory cells of the scorpion, Heterometrus fulvipes (C. Koch)

Changes in the activity of the neurosecretory cells in the cephalothoracic nerve mass (CTNM) of the scorpion, Heterometrus fulvipes (C. Koch) was studied over a 24-h period under natural light-dark cycles. Cytological and cytometric observations revealed that the first 3 groups of type 'A' cells in the sub-oesophageal ganglion of CTNM evidenced ultradian variations in their activity. Two peaks were seen in the activity of these neurosecretory cells, at 12(00) and 00(00). It is inferred that the cells show a biphasic rhythm in the synthesis of two different principles possibly having antagonistic effects on the central nervous system.     

Purification and partial characterization of an erythroagglutinin from the hemolymph of scorpion, Heterometrus bengalensis

An erythroagglutinin from the hemolymph of the scorpion, Heterometrus bengalensis, has been purified by gel filtration and ion-exchange chromatography. Its homogeneity has been demonstrated by polyacrylamide gel electrophoresis. The purified agglutinin appears to be a monomeric protein having a possible molecular weight between 146,000 and 148,000. It has no divalent cation requirement for erythroagglutination. The erythroagglutination is not inhibited by saccharides, glycoproteins and mucin. Identical erythroagglutination pattern is obtained with normal as well as neuraminidase treated erythrocytes.     

Purification and properties of phospholipase A2 from the venom of scorpion, (Heterometrus fulvipes)

Phospholipase A2 was purified about 78 fold from the venom of scorpion Heterometrus fulvipes. The molecular weight of the enzyme was found to be 16,000 and it was optimally active at pH 7.4 and at 50 degrees C. The Km value of the enzyme was 1.8 x 10(-3) M. Calcium, Magnesium and Zinc ions stimulated whereas Mercury ion and EDTA inhibited the enzyme activity. This enzyme exhibited fluorescence emission maximum between 310-320 nm.     

Purification of a unique glycoprotein that enhances phenol oxidase activity in scorpion (Heterometrus bengalensis) haemolymph.

A monomeric glycoprotein (SGP) of Mr 32,000 was isolated to purity from scorpion (Heterometrus bengalensis) haemolymph by (NH4)2SO4 fractionation, chromatofocusing and h.p.l.c. The homogeneity of SGP is confirmed by polyacrylamide-gel electrophoresis. SGP is soluble in 100%-satd. (NH4)2SO4 solution. Needle-shaped crystals of SGP were obtained in an aqueous environment. The glycan part of the molecule contains arabinose, which does not commonly occur in animal glycoproteins. Amino acid analysis demonstrated a preponderance of glycine, tyrosine and glutamic acid. SGP enhances phenol oxidase (EC 1.14.18.1) activity.     

Proteomic analysis of the venom of Heterometrus longimanus (Asian black scorpion)

Venoms have evolved over millions of years into potent cocktails of bioactive peptides and proteins. These compounds can be of great value to the pharmaceutical industry for numerous clinical applications. In this study, a novel proteomic - bioinformatic approach was utilised, where chromatography followed by gel electrophoresis was utilised to separate the venom peptides/proteins of Heterometrus longimanus (Asian black scorpion). Purified peptides were analysed by tandem mass spectrometry, de novo sequenced and then homology matched against known peptides in the Swiss-Prot protein database. Numerous potentially biologically active peptide matches were discovered, and a simple scoring system applied to putatively assign functions to the peptides. As a validation of this approach, the functional composition of the experimentally derived proteome is similar to that of other scorpions, and contains a potent mix of toxins, antimicrobials and ionic channel inhibitors.     

Neural factors in the regulation of oxygen consumption rhythm in the tissues of the scorpion, Heterometrus fulvipes (C. Koch)

Circadian variation in the effect of cephalothoracic nerve mass (CTNM) extract and blood of the oxygen consumption of the isolated hepatopancreas of the scorpion, Heterometrus fulvipes, was studied. Both CTNM extract and blood from 2000 animal cause maximum increase in oxygen consumption. Another maximum increase was also noted at 0800. It is inferred that a chemical factor from the CTNM is responsible for the observed changes in oxygen consumption of hepatopancreas.     

Isolation and characterization of the insecticidal,two-domain toxin LaIT3 from the Liocheles australasiae scorpion venom

ABSTRACT

A novel insecticidal peptide (LaIT3) was isolated from the Liocheles australasiae venom. The primary structure of LaIT3 was determined by a combination of Edman degradation and MS/MS de novo sequencing analysis. Discrimination between Leu and Ile in MS/MS analysis was achieved based on the difference in side chain fragmentation assisted by chemical derivatization. LaIT3 was determined to be an 84-residue peptide with three intrachain disulfide bonds. The sequence similarity search revealed that LaIT3 belongs to the scorpine-like peptides consisting of two structural domains: an N-terminal α-helical domain and a C-terminal cystine-stabilized domain. As observed for most of the scorpine-like peptides, LaIT3 showed significant antibacterial activity against Escherichia coli, which is likely to be caused by its membrane-disrupting property.     

Isolation and primary structure of a potent toxin from the venom of the scorpion Centruroides sculpturatus Ewing.

A potent toxin has been purified from the venom of the scorpion Centruroides sculpturatus Ewing using the ion-exchange resin CM-Sepharose CL-6B at basic pH. The toxin, designated CsE M1, comprised 65 amino acid residues and its primary structure was established as: Lys-Glu-Gly-Tyr-Leu-Val-Asn-Ser-Tyr-Thr10-Gly-Cys-Lys-Tyr-Glu-Cys- Leu-Lys-Leu- Gly20-Asp-Asn-Asp-Tyr-Cys-Leu-Arg-Glu-Cys-Arg30-Gln-Gln-Tyr- Gly-Lys-Ser-Gly-Gly - Tyr-Cys40-Tyr-Ala-Phe-Ala-Cys-Trp-Cys-Thr-His-Leu50-Tyr-Glu- Gln-Ala-Val-Val-Trp - Pro-Leu-Pro60-Asn-Lys-Thr-Cys-Asn. CsE M1 is the most lethal protein to be identified in C. sculpturatus venom and the LD50 of the toxin, determined by subcutaneous injection into Swiss mice, is 87 micrograms/kg. CsE M1 shows strong structural similarity (92% positional identity) to the most potent beta-toxin, Css II, from the Mexican scorpion, Centruroides suffusus suffusus but is quite dissimilar to the previously characterized toxins with low potency isolated from C. sculpturatus Ewing.