Isolation,structural identification and biological characterization of two conopeptides from the Conus pennaceus venom

A novel anti-mollusk conopeptide pn4c was isolated from the Conus pennaceus venom by repeated HPLC fractionation based on the activity against freshwater snails. The primary structure of pn4c was determined by the mass spectrometric de novo sequencing analysis. In addition, pn3a was isolated from the same fraction containing pn4c, as a peptide with unknown functions.     

Snake venom: From fieldwork to the clinic: Recent insights into snake biology, together with new technology allowing high-throughput screening of venom, bring new hope for drug discovery

Snake venoms are recognized here as a grossly under-explored resource in pharmacological prospecting. Discoveries in snake systematics demonstrate that former taxonomic bias in research has led to the neglect of thousands of species of potential medical use. Recent discoveries reveal an unexpectedly vast degree of variation in venom composition among snakes, from different species down to litter mates. The molecular mechanisms underlying this diversity are only beginning to be understood. However, the enormous potential that this resource represents for pharmacological prospecting is clear. New high-throughput screening systems offer greatly increased speed and efficiency in identifying and extracting therapeutically useful molecules. At the same time a global biodiversity crisis is threatening the very snake populations on which hopes for new venom-derived medications depend. Biomedical researchers, pharmacologists, clinicians, herpetologists, and conservation biologists must combine their efforts if the full potential of snake venom-derived medications is to be realized.     

Tetanus and Botulinum Toxins Inhibit, and Black Widow Spider Venom Stimulates the Release of Methionine-Enkephalin-Like Material In Vitro

The actions of tetanus toxin, botulinum A toxin, and black widow spider venom on the release of methionine-enkephalin-like immunoreactivity have been studied; a particulate fraction prepared from rat striata was used. Depending on the duration of preincubation, tetanus toxin diminished the release evoked by veratridine (50 microM final concentration), and abolished it at final concentrations between 0.1 and 1 micrograms/ml. Botulinum A toxin was about 10 to 20 times less potent. Heating or pretreatment with antitoxin inactivated the clostridial toxins. The particulate fraction pretreated with V. cholerae neuraminidase retained its toxin sensitivity. Tetanus toxin also depressed the release due to sea anemone toxin II and high K+. Spider venom stimulated the release in a concentration-dependent manner and required the presence of Ca2+; its effects were depressed by tetanus toxin. These results support the view that both clostridial toxins and spider venom act as broad-range presynaptic neurotoxins on peptidergic transmitter systems.     

Adenosine in the venoms from viperinae snakes of the genus Bitis: identification and quantitation using LC/MS and CE/MS

Snake venoms are rich sources of toxic proteins and small molecules. This study was directed at molecules of molecular mass below 1 kDa. Thirty different venoms, of either neurotoxic or haemorrhagic type, were fractionated using size-exclusion chromatography. Only venoms of the Puff adder (Bitis arietans), Gaboon viper (Bitis gabonica), and Rhinoceros viper (Bitis nasicornis) exhibited large absorbance peaks at lambda(280 nm) in the total volume range of the chromatographic column indicating the presence of abundant low molecular mass material. Analysis of fractions containing this material using both HPLC and capillary electrophoresis interfaced with electrospray ion-trap mass spectrometry unequivocally established that the bioactive nucleoside, adenosine, was the major component. The concentrations of adenosine found (Puff adder--97.7 x 10(-6) mol L(-1); Gaboon viper--28.0 x 10(-6) mol L(-1); and Rhinoceros viper-56.8 x 10(-6) mol L(-1)) were above those required to activate all known sub-types of adenosine receptors. Adenosine may thus act at the site of envenomation causing local vasodilatation and may play a role in the subsequent systemic hypotension observed.     

Growth and protein phosphorylation in the Nb2 lymphoma: effect of prolactin, cAMP, and agents that activate adenylate cyclase

The Nb2 T lymphoma is unique in that these lymphocytes proliferate in response to prolactin as well as in response to interleukin-2. In this study, we have examined the responsiveness of the adenylate cyclase system in Nb2 cells and the role of this signaling system in regulating proliferation and protein phosphorylation. An analog of cAMP inhibited prolactin-stimulated proliferation and blocked a prolactin-induced decrease in protein phosphorylation. Forskolin, a potent activator of adenylate cyclase in T lymphocytes, did not elevate cAMP levels in Nb2 cells and was not an effective inhibitor of prolactin-induced proliferation. In fact, one preparation of forskolin stimulated proliferation of quiescent Nb2 cells. Like forskolin, prostaglandin E2 did not stimulate cAMP production in Nb2 cells even though it increased cAMP in a preparation of rat peripheral blood lymphocytes. Cholera toxin appeared to ADP-ribosylate a stimulatory guanine nucleotide-binding protein in Nb2 cells, but the toxin did not increase intracellular levels of cAMP nor was it a potent anti-mitogenic agent. Pertussis toxin, an agent that can increase cAMP production through suppression of the inhibitory guanine nucleotide-binding protein, exerted only minor anti-proliferative actions on prolactin-stimulated Nb2 cells. These data suggest that cAMP inhibits Nb2 cell proliferation and prolactin-induced changes in protein phosphorylation but that the adenylate cyclase system in our clone of Nb2 cells responds poorly to agents that normally increase cAMP.     

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.     

Field evaluation of alpha-chlorohydrin against the Indian mole rat: studies on toxic and antifertility effects

Field acceptance and efficacy of a toxicant-sterilant, alpha-chlorohydrin (α-CH), at its 0.5% concentration in bait were evaluated against rodents in sugarcane fields which harboured high populations of the Indian mole rat Bandicota bengalensis (Trap Index (TI) = 53.6 rats/100 traps/24 h) followed by that of the Indian bush rat Golunda ellioti (TI = 28.6) and soft-furred field rat Rattus meltada (TI = 1.8). Acceptance of the α-CH bait by rodents in the fields was evident from complete consumption of the offered bait at most of the baiting points. The treatment (72 h exposure to the poison bait) resulted in 63.7% to 82.9% rodent mortality. Survey of the mature sugarcane crop revealed that the percentage of rodent cut canes in the treated fields (7.6% to 16.2%) was significantly less than that of the reference fields (26.3%). Most of the surviving male B. bengalensis, captured after 15 days of the treatment, had developed sterility as revealed by the functional abnormalities in their testes and epid-idymides. They showed decreased thickness of the seminiferous tubules, lower population of spermatogenic cells, cauda epididymal sperm concentration, live sperm and sperm mortality. A nonsignificant positive correlation between the testicular weight and sperm motility in males from treated fields indicated the effect of α-CH at maturation level in the cauda epididymides. Overall, it was evident that a significant proportion of the surviving B. bengalensis had become permanently sterile as a consequence to the development of spermatocoeles in the caput epididymides and the remaining had significantly low numbers (< 30%) of motile sperm in their cauda epididymides which was, obviously, a handicap for successful fertilisation.     

Crystallographic and spectroscopic characterization of a molecular hinge: Conformational changes in bothropstoxin I,a dimeric Lys49-phospholipase A2 homologue

Bothropstoxin I (BthTX-I) from the venom of Bothrops jararacussuis a myotoxic phospholipase A2 (PLA2) homologue which, although catalytically inactive due to an Asp49→Lys substitution, disrupts the integrity of lipid membranes by a Ca²⁺-independent mechanism. The crystal structures of two dimeric forms of BthTX-I which diffract X-rays to resolutions of 3.1 and 2.1 Å have been determined. The monomers in both structures are related by an almost perfect twofold axis of rotation and the dimer interfaces are defined by contacts between the N-terminal α-helical regions and the tips of the β-wings of partner monomers. Significant differences in the relative orientation of the monomers in the two crystal forms results in “open” and “closed” dimer conformations. Spectroscopic investigations of BthTX-I in solution have correlated these conformational differences with changes in the intrinsic fluorescence emission of the single tryptophan residues located at the dimer interface. The possible relevance of this structural transition in the Ca²⁺-independent membrane damaging activity is discussed. Proteins 30:442–454, 1998. © 1998 Wiley-Liss, Inc.     

And you thought we had moved beyond all that: biological race returns to the social sciences

Recently, sociologists have argued in high-profile journals that racial categories are linked to genetically distinct clusters within the human population. They propose theorizing race as a socially constructed categorization system that is related to biological groupings within our species. This work overlooks, however, the extent to which statistically inferred genetic clusters are themselves socially constructed, making it impossible to juxtapose ‘subjective’ social categories with ‘objective’ biological ones. This editorial urges social scientists to take a critical look at claims about the genetic underpinnings of race, and to contribute their insights to ongoing debates about the nature of race.     

Logical puzzles and scientific controversies: The nature of species,viruses and living organisms

In the past, biologists believed that species were stable and permanent entities and they viewed them as natural kinds which, like the chemical elements, exist in nature independently of any human conceptualization. After Darwin, biologists came to accept that species were the products of evolution and natural selection and were not immutable natural kinds.     

Identification, cloning and expression of a second gene (vpr1) from the venom of the endoparasitic wasp, Pimpla hypochondriaca that displays immunosuppressive activity

Previously, we biochemically isolated an immunosuppressive protein (VPr3) from the venom of Pimpla hypochondriaca and cloned and expressed the gene in bacteria. The deduced amino acid sequence for VPr3 shares 63% identity with a second P. hypochondriaca protein, venom protein one (VPr1). We have now cloned and expressed the gene for vpr1. The expression of His-tagged recombinant VPr1 (rVPr1) in E. coli BL21 Star™ (DE3) cells was induced by the addition of 0.5 mM IPTG. Cultures were grown at 24 and 37 °C, and VPr1 more readily partitioned into the soluble fraction at 24 °C. Soluble rVPr1 was purified using the MagneHis purification system and a modified elution buffer to allow the protein to be directly tested for activity against haemocytes. It was observed that rVPr1 prevented the ability of haemocytes to spread and form aggregates in vitro in a dose-dependent manner. Furthermore, comparable levels of activity were observed when similar concentrations of rVPr1 and rVPr3 were tested. In addition, the encapsulation of Sephadex beads in vivo was reduced by the presence of rVPr1 and beads were unencapsulated (negative) or only weakly encapsulated. The functional and physio-chemical properties of rVPr1 and rVPr3 are compared and discussed.     

Arundifungin, a novel antifungal compound produced by fungi: biological activity and taxonomy of the producing organisms

Echinocandins, the lipopeptide class of glucan synthase inhibitors, are an alternative to ergosterol-synthesis inhibitors to treat candidiasis and aspergillosis. Their oral absorption, however, is low and they can only be used parenterally. During a natural product screening program for novel types of glucan synthesis inhibitors with improved bioavailability, a fungal extract was found that inhibited the growth of both a wild-type Saccharomyces cerevisiae strain and the null mutant of the FKS1 gene (fks1::HIS). The mutant strain was more sensitive to growth inhibition, suggesting that the fungal extract could contain an inhibitor of glucan synthesis. A novel acidic steroid, named arundifungin, was purified from a fungal extract obtained from a liquid culture of Arthrinium arundinis collected in Costa Rica. Arundifungin caused the same pattern of hallmark morphological alterations in Aspergillus fumigatus hyphae as echinocandins, further supporting the idea that arundifungin belongs to a new class of glucan synthesis inhibitors. Moreover, its antifungal spectrum was comparable to those of echinocandins and papulacandins, preferentially inhibiting the growth of Candida and Aspergillus strains, with very poor activity against Cryptococcus. Arundifungin was also detected in nine other fungal isolates which were ecologically and taxonomically unrelated, as assessed by sequencing of the ITS1 region. Further, it was also found in two more Arthrinium spp from tropical and temperate regions, in five psychrotolerant conspecific isolates collected on Macquarie Island (South Pacific) and belonging to the Leotiales, and in two endophytes collected in central Spain (a sterile fungus belonging to the Leotiales and an undetermined coelomycete).     

Human-Spider Entanglements: Understanding and Managing the Good,the Bad,and the Venomous

ABSTRACT

Considering the fear that spiders can generate in humans, examining human-spider interactions in urban settings may at first glance appear odd. However, human-spider interactions, which occur quite frequently in urban settings, do not necessarily have to be negative; they can, in some cases, foster respect and tolerance (sometimes through avoidance). When one considers how global transformations, invasive species, urbanization, and adaptation will impact human-spider interactions, a review of the literature pertaining to these encounters is timely. We begin this discussion by describing spiders and providing an overview of some of their positive and negative impacts. Challenges regarding species identification and envenoming are also discussed. After the role of biological, psychological, and social aspects in human-spider interactions are re-examined, we provide future options aimed at organizing broad-scale public programs for five specific target groups: 1) the general public, 2) health professionals, 3) educators, 4) naturalists, and 5) researchers. In the conclusion, we provide potential management and educational strategies aimed at increasing our knowledge and tolerance of these animals in urban settings.     

The present status of biological effects of toxic metals in the environment: lead, cadmium, and manganese

The number of reports concerning the chemical toxicology of metals which are released in the environment by natural as well as anthropogenic sources, have been increasing constantly. Lead, cadmium, and manganese have found a variety of uses in industry, craft, and agriculture owing to their physical and chemical properties. The environmental burden of heavy metals has been rising substantially by smelter emission in air and waste sewage in water. Further, organic compounds of lead and manganese used as antiknock substances in gasoline are emitted into the atmosphere by automobile exhaustion. Such environmental contamination of air, water, soil, and food is a serious threat to all living kinds. Although these metals are known to produce their toxic effects on a variety of body systems, much emphasis has been placed on their effects on the nervous system owing to apparent association of relatively low or "subclinical" levels of metallic exposure with behavioral and psychological disorders. Clinical and animal data on environmental exposure show that while lead and manganese are most toxic to the nervous system, cadmium exerts profound adverse effects on kidney and the male reproductive system. It appears that the consequences of exposure to lead in adults are less severe than the types of exposure associated with hyperactivity in neonates. Except for a few reports, hyperactivity has indeed been observed in animals exposed to either of these three metals. Experimental work has also shown that these metals produce behavioral changes by altering the metabolism of brain neurotransmitters, especially catecholamines. Recently, it is hypothesized that these metals exert their toxic effect by damaging biological defences which exist in the body to serve as protective mechanisms against exogenous toxins. A voluminous publication list with diverse opinions on the biological effects of metals is available and there is an urgent need to compile assessment of the existing literature to identify the future theme of research work. The problem of metal toxicity becomes even more complex owing to simultaneous or successive exposure of the general population to different physical, chemical, biological, and psychological factors in the environment. The net toxic manifestations produced by multiple exposure should, therefore, be different from those produced by a single factor as the result of their additive, synergistic or antagonistic action. Even though a metal may not exist in sufficient amounts to cause any disability, the toxicity could result when a second factor is also present.(ABSTRACT TRUNCATED AT 400 WORDS)     

Properties and action mechanism of the toxic lectin modeccin: Interaction with cell lines resistant to modeccin,abrin, and ricin

The toxic lectin modeccin, which inhibits protein synthesis in eukaryotic cells, is cleaved upon treatment with 2-mercaptoethanol into two peptide chains which move in polyacrylamide gels at rates corresponding to molecular weights 28,000 and 38,000. After reduction, the toxin loses its effect on cells, while its ability to inhibit cell-free protein synthesis increases. Like abrin and ricin it inhibits protein synthesis by inactivating the 60S ribosomal subunits. Modeccin binds to surface receptors containing terminal galactose residues. Competition experiments with various glycoproteins indicate that the modeccin receptors are different from the abrin receptors. In addition, they were present on HeLa cells in much smaller numbers. Moreover, mutant lines resistant to abrin and ricin were not resistant to modeccin and vice-versa. The toxin resistance of various mutant cell lines could not be accounted for by a reduced number of binding sites on cells. The data are consistent with the view that the cells possesss different populations of binding sites with differences in ability to facilitate the uptake of the toxins and that in the resistant lines the most active receptors have been reduced or eliminated.     

Phylogenetic analysis of ascomycete yeasts that form coenzyme Q-9 and the proposal of the new genera Babjeviella, Meyerozyma, Millerozyma, Priceomyces, and Scheffersomyces

Species assigned to the genera Debaryomyces, Lodderomyces, Spathaspora, and Yamadazyma, as well as selected species of Pichia and Candida that also form coenzyme Q-9, were phylogenetically analyzed from the combined sequences of the D1/D2 domains of the large subunit and the nearly complete small subunit rRNA genes. Species assigned to Debaryomyces partitioned into three clades and species assigned to Pichia were distributed among six clades. These well-supported clades were interpreted as genera, and from this analysis, the following new genera are proposed: Babjeviella, Meyerozyma, Millerozyma, Priceomyces, and Scheffersomyces. The genus Schwanniomyces was reinstated and emended, and the genus Yamadazyma was phylogenetically defined. From this study, 23 new combinations and 3 new ranks are proposed. The preceding genera are members of a single, large clade, and it is proposed to delineate this clade as the new family Debaryomycetaceae.     

Model of the toxic complex of anthrax: responsive conformational changes in both the lethal factor and the protective antigen heptamer

The toxic complex of anthrax is formed when the monomeric protective antigen (PA) (83 kDa), while bound to its cell-surface receptor, is first converted to PA63 heptamers (PA63h) following N-terminal proteolytic cleavage, and then lethal (LF) (90 kDa) or edema factor (EF) binds to the heptamer. We report a "pseudoatomic" model for the complex of PA63h and full-length LF determined by applying the normal-mode flexible fitting procedure to a approximately 18 A cryo-electron microscopy (EM) density map of the complex. The model describes the interacting surface that buries a total area of approximately 10,140 A2 comprising approximately 40% charged, and approximately 30% each of polar and hydrophobic residues. For the heptamer, the buried surface, composed of approximately 110 residues, involves primarily three monomers and includes for two, similar stretches of the polypeptide chain from domain 1. For LF, the interface again involves approximately 110 residues, mostly from the N-terminal domain I (LF(N)), and the structurally homologous C-terminal domain IV. Most interestingly, bound LF displays a marked conformational change resulting from a "collapse" of domains I, III, and IV on domain II, with the largest movement of approximately 9 A noted for domain I. On the other hand, primarily, rigid-body movements, larger than approximately 10 A for three PA63 monomers, cause the hourglass-shaped heptamer lumen to enlarge by as much as approximately 50% near the middle of the molecule. Such concerted structural rearrangements in LF and the heptamer can facilitate ingress of the ligand into the heptamer lumen prior to unfolding and release through the PA63h channel formed in the acidic late endosomal membrane.     

云南锦斑蛾幼虫体表分泌物氰苷类成分的分离与鉴定及其对黑头酸臭蚁的生物活性

为研究云南锦斑蛾Achelura yunnanensis幼虫的化学防御策略, 利用硅胶柱色谱和HPLC制备色谱等色谱学方法对其毒性分泌液进行了化学成分的分离, 并通过核磁共振和质谱学方法对分离到的成分进行了结构鉴定。从其毒性分泌液中分离得到了两个神经毒性氰苷类化合物, 经鉴定分别为linamarin和lotaustralin。取食试验表明, linamarin对黑头酸臭蚁Tapinoma melanocephalum有明显的拒食活性。我们推测, 云南锦斑蛾体内的神经毒性物质氰苷是通过摄取宿主植物冬樱花Prunus cerasoides和云南樱花P. majestic而获得的, 并在体内转化形成毒液, 用于防御其天敌。本研究为云南锦斑蛾和宿主植物的协同进化提供了化学依据。