RESUMEN
The molecular origin of waglerin peptides has remained enigmatic despite their industrial application in skin cream products to paralyse facial muscles and thus reduce the incidence of wrinkles. Here we show that these neurotoxic peptides are the result of de novo evolution within the prepro region of the C-type natriuretic peptide gene in Tropidolaemus venoms, at a site distinct from the domain encoding for the natriuretic peptide. It is the same region that yielded the azemiopsin peptides from Azemiops feae, indicative of a close relationship of this toxin gene between these two genera. The precursor region for the molecular evolution is a biodiversity hotspot that has yielded other novel bioactive peptides with novel activities. We detail the diversity of components in this and other species in order to explore what characteristics enable it to be such a biodiscovery treasure trove. The unusual function of Tropidolaemus venoms may have been selected for due to evolutionary pressures brought about by a high likelihood of prey escape.
Asunto(s)
Venenos de Crotálidos/genética , Venenos de Víboras/uso terapéutico , Secuencia de Aminoácidos/genética , Animales , Evolución Biológica , Venenos de Crotálidos/uso terapéutico , Venenos de Crotálidos/toxicidad , Evolución Molecular , Datos de Secuencia Molecular , Péptidos/química , Filogenia , Crema para la Piel , Venenos de Víboras/toxicidadRESUMEN
Marine cone snail venoms are highly complex mixtures of peptides and proteins. They have been studied in-depth over the past 3 decades, but the modus operandi of the venomous apparatus still remains unclear. Using the fish-hunting Conus consors as a model, we present an integrative venomics approach, based on new proteomic results from the venom gland and data previously obtained from the transcriptome and the injectable venom. We describe here the complete peptide content of the dissected venom by the identification of numerous new peptides using nanospray tandem mass spectrometry in combination with transcriptomic data. Results reveal extensive mature peptide diversification mechanisms at work in the venom gland. In addition, by integrating data from three different venom stages, transcriptome, dissected, and injectable venoms, from a single species, we obtain a global overview of the venom processing that occurs from the venom gland tissue to the venom delivery step. In the light of the successive steps in this venom production system, we demonstrate that each venom compartment is highly specific in terms of peptide and protein content. Moreover, the integrated investigative approach discussed here could become an essential part of pharmaceutical development, as it provides new potential drug candidates and opens the door to numerous analogues generated by the very mechanisms used by nature to diversify its peptide and protein arsenal.
Asunto(s)
Conotoxinas/toxicidad , Caracol Conus/química , Proteómica/métodos , Secuencia de Aminoácidos , Animales , Cromatografía Líquida de Alta Presión , Espectrometría de Masas , Datos de Secuencia Molecular , Homología de Secuencia de AminoácidoRESUMEN
Conotoxins are venom peptides from cone snails with multiple disulfide bridges that provide a rigid structural scaffold. Typically acting on ion channels implicated in neurotransmission, conotoxins are of interest both as tools for pharmacological studies and as potential new medicines. δ-Conotoxins act by inhibiting inactivation of voltage-gated sodium channels (Nav). Their pharmacology has not been extensively studied because their highly hydrophobic character makes them difficult targets for chemical synthesis. Here we adopted an acid-cleavable solubility tag strategy that facilitated synthesis, purification, and directed disulfide bridge formation. Using this approach we readily produced three native δ-conotoxins from Conus consors plus two rationally designed hybrid peptides. We observed striking differences in Nav subtype selectivity across this group of compounds, which differ in primary structure at only three positions: 12, 23, and 25. Our results provide new insights into the structure-activity relationships underlying the Nav subtype selectivity of δ-conotoxins. Use of the acid-cleavable solubility tag strategy should facilitate synthesis of other hydrophobic peptides with complex disulfide bridge patterns.
Asunto(s)
Conotoxinas/síntesis química , Activación del Canal Iónico/fisiología , Fragmentos de Péptidos/síntesis química , Canales de Sodio Activados por Voltaje/fisiología , Ácidos/química , Secuencia de Aminoácidos , Animales , Cromatografía Líquida de Alta Presión , Conotoxinas/química , Conotoxinas/farmacología , Caracol Conus/química , Disulfuros/química , Relación Dosis-Respuesta a Droga , Femenino , Activación del Canal Iónico/efectos de los fármacos , Activación del Canal Iónico/genética , Datos de Secuencia Molecular , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Oocitos/fisiología , Fragmentos de Péptidos/química , Fragmentos de Péptidos/farmacología , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiología , Solubilidad , Espectrometría de Masa por Ionización de Electrospray , Relación Estructura-Actividad , Canales de Sodio Activados por Voltaje/genética , Xenopus laevisRESUMEN
The glycopeptide CcTx, isolated from the venom of the piscivorous cone snail Conus consors, belongs to the κA-family of conopeptides. These toxins elicit excitotoxic responses in the prey by acting on voltage-gated sodium channels. The structure of CcTx, a first in the κA-family, has been determined by high-resolution NMR spectroscopy together with the analysis of its O-glycan at Ser7. A new type of glycopeptide O-glycan core structure, here registered as core typeâ 9, containing two terminal L-galactose units {α-L-Galp-(1â4)-α-D-GlcpNAc-(1â6)-[α-L-Galp-(1â2)-ß-D-Galp-(1â3)-]α-D-GalpNAc-(1âO)}, is highlighted. A sequence comparison to other putative members of the κA-family suggests that O-linked glycosylation might be more common than previously thought. This observation alone underlines the requirement for more careful and in-depth investigations into this type of post-translational modification in conotoxins.
Asunto(s)
Caracol Conus/química , Glicopéptidos/química , Venenos de Moluscos/química , Animales , Glicosilación , Espectroscopía de Resonancia Magnética , Estructura MolecularRESUMEN
Cone snail venoms are considered an untapped reservoir of extremely diverse peptides, named conopeptides, displaying a wide array of pharmacological activities. We report here for the first time, the presence of high molecular weight compounds that participate in the envenomation cocktail used by these marine snails. Using a combination of proteomic and transcriptomic approaches, we identified glycosyl hydrolase proteins, of the hyaluronidase type (Hyal), from the dissected and injectable venoms ("injectable venom" stands for the venom variety obtained by milking of the snails. This is in contrast to the "dissected venom", which was obtained from dissected snails by extraction of the venom glands) of a fish-hunting cone snail, Conus consors (Pionoconus clade). The major Hyal isoform, Conohyal-Cn1, is expressed as a mixture of numerous glycosylated proteins in the 50 kDa molecular mass range, as observed in 2D gel and mass spectrometry analyses. Further proteomic analysis and venom duct mRNA sequencing allowed full sequence determination. Additionally, unambiguous segment location of at least three glycosylation sites could be determined, with glycans corresponding to multiple hexose (Hex) and N-acetylhexosamine (HexNAc) moieties. With respect to other known Hyals, Conohyal-Cn1 clearly belongs to the hydrolase-type of Hyals, with strictly conserved consensus catalytic donor and positioning residues. Potent biological activity of the native Conohyals could be confirmed in degrading hyaluronic acid. A similar Hyal sequence was also found in the venom duct transcriptome of C. adamsonii (Textilia clade), implying a possible widespread recruitment of this enzyme family in fish-hunting cone snail venoms. These results provide the first detailed Hyal sequence characterized from a cone snail venom, and to a larger extent in the Mollusca phylum, thus extending our knowledge on this protein family and its evolutionary selection in marine snail venoms.
Asunto(s)
Caracol Conus/enzimología , Glicósido Hidrolasas/metabolismo , Venenos de Moluscos/enzimología , Secuencia de Aminoácidos , Animales , Caracol Conus/metabolismo , Perfilación de la Expresión Génica , Glicósido Hidrolasas/química , Glicosilación , Hialuronoglucosaminidasa/química , Hialuronoglucosaminidasa/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Peso Molecular , Venenos de Moluscos/metabolismo , N-Glicosil Hidrolasas/química , N-Glicosil Hidrolasas/metabolismo , Filogenia , Estructura Secundaria de Proteína , Proteómica/métodos , ARN Mensajero/metabolismo , Homología de Secuencia de AminoácidoRESUMEN
Major variations in venom composition can occur between juvenile and adult venomous snakes. However, due to logistical constraints, antivenoms are produced using adult venoms in immunising mixtures, possibly resulting in limited neutralisation of juvenile snake venoms. Daboia russelii is one of the leading causes of snakebite death across South Asia. Its venom is potently procoagulant, causing stroke in prey animals but causing in humans consumptive coagulopathy-a net anticoagulant state-and sometimes death resulting from hemorrhage. In this in vitro study, we compared the venom activity of-and antivenom efficacy against-six 2-week-old D. russelii relative to that of their parents. Using a coagulation analyser, we quantified the relative coagulotoxicity of these venoms in human, avian, and amphibian plasma. The overall potency on human plasma was similar across all adult and neonate venoms, and SII (Serum Institute of India) antivenom was equipotent in neutralising these coagulotoxic effects. In addition, all venoms were also similar in their action upon avian plasma. In contrast, the neonate venoms were more potent on amphibian plasma, suggesting amphibians make up a larger proportion of neonate diet than adult diet. A similar venom potency in human and avian plasmas but varying selectivity for amphibian plasma suggests ontogenetic differences in toxin isoforms within the factor X or factor V activating classes, thereby providing a testable hypothesis for future transcriptomics work. By providing insights into the functional venom differences between adult and neonate D. russelii venoms, we hope to inform clinical treatment of patients envenomated by this deadly species and to shed new light on the natural history of these extremely medically important snakes.
Asunto(s)
Daboia , Mordeduras de Serpientes , Animales , Antivenenos/farmacología , Humanos , Recién Nacido , Serpientes , PonzoñasRESUMEN
Bites from helodermatid lizards can cause pain, paresthesia, paralysis, and tachycardia, as well as other symptoms consistent with neurotoxicity. Furthermore, in vitro studies have shown that Heloderma horridum venom inhibits ion flux and blocks the electrical stimulation of skeletal muscles. Helodermatids have long been considered the only venomous lizards, but a large body of robust evidence has demonstrated venom to be a basal trait of Anguimorpha. This clade includes varanid lizards, whose bites have been reported to cause anticoagulation, pain, and occasionally paralysis and tachycardia. Despite the evolutionary novelty of these lizard venoms, their neuromuscular targets have yet to be identified, even for the iconic helodermatid lizards. Therefore, to fill this knowledge gap, the venoms of three Heloderma species (H. exasperatum, H. horridum and H. suspectum) and two Varanus species (V. salvadorii and V. varius) were investigated using Gallus gallus chick biventer cervicis nerve-muscle preparations and biolayer interferometry assays for binding to mammalian ion channels. Incubation with Heloderma venoms caused the reduction in nerve-mediated muscle twitches post initial response of avian skeletal muscle tissue preparation assays suggesting voltage-gated sodium (NaV) channel binding. Congruent with the flaccid paralysis inducing blockage of electrical stimulation in the skeletal muscle preparations, the biolayer interferometry tests with Heloderma suspectum venom revealed binding to the S3-S4 loop within voltage-sensing domain IV of the skeletal muscle channel subtype, NaV1.4. Consistent with tachycardia reported in clinical cases, the venom also bound to voltage-sensing domain IV of the cardiac smooth muscle calcium channel, CaV1.2. While Varanus varius venom did not have discernable effects in the avian tissue preparation assay at the concentration tested, in the biointerferometry assay both V. varius and V. salvadorii bound to voltage-sensing domain IV of both NaV1.4 and CaV1.2, similar to H. suspectum venom. The ability of varanid venoms to bind to mammalian ion channels but not to the avian tissue preparation suggests prey-selective actions, as did the differential potency within the Heloderma venoms for avian versus mammalian pathophysiological targets. This study thus presents the detailed characterization of Heloderma venom ion channel neurotoxicity and offers the first evidence of varanid lizard venom neurotoxicity. In addition, the data not only provide information useful to understanding the clinical effects produced by envenomations, but also reveal their utility as physiological probes, and underscore the potential utility of neglected venomous lineages in the drug design and development pipeline.
Asunto(s)
Canales de Calcio/metabolismo , Lagartos , Neurotoxinas/toxicidad , Canales de Sodio/metabolismo , Ponzoñas/toxicidad , Animales , Pollos , Técnicas In Vitro , Masculino , Unión Neuromuscular/efectos de los fármacos , Unión Neuromuscular/fisiología , Unión ProteicaRESUMEN
Venom research is a highly multidisciplinary field that involves multiple subfields of biology, informatics, pharmacology, medicine, and other areas. These different research facets are often technologically challenging and pursued by different teams lacking connection with each other. This lack of coordination hampers the full development of venom investigation and applications. The COST Action CA19144-European Venom Network was recently launched to promote synergistic interactions among different stakeholders and foster venom research at the European level.
Asunto(s)
PonzoñasRESUMEN
Does the venom of Trimeresurus albolabris (white-lipped pit viper) differ between neonate and adults? This species is responsible for most snakebites within south and southeast Asia, yet it is unknown whether ontogenetic variation in venom composition occurs in this species, or how this might affect antivenom efficacy. Using a coagulation analyser robot, we examined the anticoagulant activity of T. albolabris venom from eight individuals across multiple age classes. We then compared the efficacy of Thai Red Cross Green Pit Viper Antivenom across these age classes. Venoms from all age classes were equally potent in their pseudo-procoagulant, fibrinogenolytic activity, in that fibrinogen was cleaved to form weak, unstable fibrin clots that rapidly broke down, thus resulting in a net anticoagulant state. Similarly, this coagulotoxic activity was well neutralised by antivenom across all venoms. Given that coagulotoxicity is the primary serious pathology in T. albolabris envenomations, we conclude that Thai Red Cross Green Tree Pit Viper Antivenom is a valid treatment for envenomations by this species, regardless of age or sex of the offending snake. These results are relevant for clinical treatment of envenomations by T. albolabris.
Asunto(s)
Coagulación Sanguínea/efectos de los fármacos , Venenos de Crotálidos/toxicidad , Mordeduras de Serpientes/terapia , Trimeresurus/fisiología , Envejecimiento , Animales , Antivenenos , Femenino , Humanos , MasculinoRESUMEN
Animal venoms are rich in hundreds of toxins with extraordinary biological activities. Their exploitation is difficult due to their complexity and the small quantities of venom available from most venomous species. We developed a Venomics approach combining transcriptomic and proteomic characterization of 191 species and identified 20,206 venom toxin sequences. Two complementary production strategies based on solid-phase synthesis and recombinant expression in Escherichia coli generated a physical bank of 3597 toxins. Screened on hMC4R, this bank gave an incredible hit rate of 8%. Here, we focus on two novel toxins: N-TRTX-Preg1a, exhibiting an inhibitory cystine knot (ICK) motif, and N-BUTX-Ptr1a, a short scorpion-CSαß structure. Neither N-TRTX-Preg1a nor N-BUTX-Ptr1a affects ion channels, the known targets of their toxin scaffolds, but binds to four melanocortin receptors with low micromolar affinities and activates the hMC1R/Gs pathway. Phylogenetically, these two toxins form new groups within their respective families and represent novel hMC1R agonists, structurally unrelated to the natural agonists.
Asunto(s)
Proteómica/métodos , Receptores de Melanocortina/agonistas , Venenos de Escorpión/farmacología , Secuencia de Aminoácidos , Animales , Células HEK293 , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Receptores de Melanocortina/metabolismo , Venenos de Escorpión/genética , Venenos de Escorpión/aislamiento & purificación , Venenos de Escorpión/metabolismoRESUMEN
The functional activities of Anguimorpha lizard venoms have received less attention compared to serpent lineages. Bite victims of varanid lizards often report persistent bleeding exceeding that expected for the mechanical damage of the bite. Research to date has identified the blockage of platelet aggregation as one bleeding-inducing activity, and destructive cleavage of fibrinogen as another. However, the ability of the venoms to prevent clot formation has not been directly investigated. Using a thromboelastograph (TEG5000), clot strength was measured after incubating human fibrinogen with Heloderma and Varanus lizard venoms. Clot strengths were found to be highly variable, with the most potent effects produced by incubation with Varanus venoms from the Odatria and Euprepriosaurus clades. The most fibrinogenolytically active venoms belonged to arboreal species and therefore prey escape potential is likely a strong evolutionary selection pressure. The results are also consistent with reports of profusive bleeding from bites from other notably fibrinogenolytic species, such as V. giganteus. Our results provide evidence in favour of the predatory role of venom in varanid lizards, thus shedding light on the evolution of venom in reptiles and revealing potential new sources of bioactive molecules useful as lead compounds in drug design and development.
Asunto(s)
Fibrinógeno/química , Lagartos , Ponzoñas/química , Animales , Coagulación Sanguínea , Humanos , TromboelastografíaRESUMEN
Piperacillin, a potent beta-lactam antibiotic, is effective in a large variety of Gram+ and Gram- bacterial infections but its administration is limited to the parenteral route as it is not absorbed when given orally. In an attempt to overcome this problem, we have synthesized a novel series of charged liposaccharide complexes of piperacillin comprising a sugar moiety derived from d-glucose conjugated to a lipoamino acid residue with varying side-chain length (cationic entity) and the piperacillin anion. A complete multiple reaction monitoring LC-MS/MS method was developed to detect and characterize the synthesized complexes. The same method was then successfully applied to assess the in vitro apparent permeability values of the charged liposaccharide complexes in Caco-2 monolayers.
Asunto(s)
Antibacterianos/farmacocinética , Cromatografía Liquida/métodos , Piperacilina/farmacocinética , Espectrometría de Masas en Tándem/métodos , Administración Oral , Antibacterianos/administración & dosificación , Células CACO-2 , Excipientes/química , Humanos , Lipopolisacáridos/química , Permeabilidad , Piperacilina/administración & dosificaciónRESUMEN
Ability of N,N'-linked oligoureas containing proteinogenic side chains to adopt a stable helix conformation in solution has been described recently. NMR as well as circular dichroism (CD) spectroscopies were employed to gain insight into their specific fold. It is herein proposed to extend the structural information available on these peptidomimetics by an advantageous use of a methylene spin state selective NMR experiment. Homodecoupling provided by the pulse scheme made it possible to readily measure conformation-dependent (3)J(HH) constants that are difficult if not impossible to obtain with standard NMR experiments. Adding those couplings to the NMR restraints improved the quality of the structure calculations significantly, as judged by a ca 30% decrease of the root mean square deviation (RMSD) obtained over an ensemble of 20 structures. Moreover, accurate determination of individual (1)J(CH) couplings within each methylene group revealed uniform values throughout the oligourea sequence, with (1)J(CH) systematically slightly larger for the pro-S hydrogen than for the pro-R. As shown in this study, the methylene spin state selective NMR experiment displays a good intrinsic sensitivity and could therefore provide valuable structural information at (13)C natural abundance for peptidomimetic molecules and foldamers bearing diastereotopic methylene protons.
Asunto(s)
Aminoácidos/química , Espectroscopía de Resonancia Magnética/métodos , Espectroscopía de Resonancia Magnética/normas , Péptidos/química , Urea/análogos & derivados , Urea/análisis , Isótopos de Carbono , Modelos Moleculares , Conformación Molecular , Pliegue de Proteína , Protones , Estándares de Referencia , Sensibilidad y Especificidad , Soluciones/química , Estereoisomerismo , Urea/síntesis químicaRESUMEN
Atractaspis snake species are enigmatic in their natural history, and venom effects are correspondingly poorly described. Clinical reports are scarce but bites have been described as causing severe hypertension, profound local tissue damage leading to amputation, and deaths are on record. Clinical descriptions have largely concentrated upon tissue effects, and research efforts have focused upon the blood-pressure affecting sarafotoxins. However, coagulation disturbances suggestive of procoagulant functions have been reported in some clinical cases, yet this aspect has been uninvestigated. We used a suite of assays to investigate the coagulotoxic effects of venoms from six different Atractaspis specimens from central Africa. The procoagulant function of factor X activation was revealed, as was the pseudo-procoagulant function of direct cleavage of fibrinogen into weak clots. The relative neutralization efficacy of South African Antivenom Producer's antivenoms on Atractaspis venoms was boomslang>>>polyvalent>saw-scaled viper. While the boomslang antivenom was the most effective on Atractaspis venoms, the ability to neutralize the most potent Atractaspis species in this study was up to 4-6 times less effective than boomslang antivenom neutralizes boomslang venom. Therefore, while these results suggest cross-reactivity of boomslang antivenom with the unexpectedly potent coagulotoxic effects of Atractaspis venoms, a considerable amount of this rare antivenom may be needed. This report thus reveals potent venom actions upon blood coagulation that may lead to severe clinical effects with limited management strategies.
Asunto(s)
Aletinofidios , Antivenenos/farmacología , Venenos de Abeja/farmacología , Trastornos de la Coagulación Sanguínea/prevención & control , Factor X/metabolismo , Factor Xa/efectos de los fármacos , África Central , Animales , Especificidad de Anticuerpos , Coagulación Sanguínea/efectos de los fármacos , Trastornos de la Coagulación Sanguínea/inducido químicamente , Reacciones Cruzadas , Fibrinógeno/efectos de los fármacos , Humanos , Técnicas In Vitro , TromboelastografíaRESUMEN
Unnatural oligomeric scaffolds designed to adopt defined secondary structures (e.g., helices), while retaining the chemical diversity of amino acid side chains, are of practical value to elaborate functional mimetics of bioactive alpha-polypeptides. Enantiopure N,N'-linked oligoureas as short as seven residues long have been previously shown to fold into a stable helical structure, stabilized by 12- and 14-membered H-bonded rings. We now report that eight-residue oligoureas designed to mimic globally amphiphilic alpha-helical host-defense peptides are effective against both gram-negative and gram-positive bacteria (including methicillin-resistant Staphylococcus aureus [MRSA]) and exhibit selectivity for bacterial versus mammalian cells. Circular dichroism (CD) spectroscopy studies suggest enhanced helical propensity of oligoureas in the presence of phospholipid vesicles. The utility of this new class of nonpeptidic foldamers for biological applications is highlighted by high resistance to proteolytic degradation.
Asunto(s)
Antibacterianos/química , Biopolímeros , Imitación Molecular , Péptidos/química , Secuencia de Aminoácidos , Cromatografía Líquida de Alta Presión , Dicroismo Circular , Resistencia a la Meticilina , Estructura Secundaria de Proteína , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Staphylococcus aureus/efectos de los fármacosRESUMEN
While snake venoms have been the subject of intense study, comparatively little work has been done on lizard venoms. In this study, we have examined the structural and functional diversification of anguimorph lizard venoms and associated toxins, and related these results to dentition and predatory ecology. Venom composition was shown to be highly variable across the 20 species of Heloderma, Lanthanotus, and Varanus included in our study. While kallikrein enzymes were ubiquitous, they were also a particularly multifunctional toxin type, with differential activities on enzyme substrates and also ability to degrade alpha or beta chains of fibrinogen that reflects structural variability. Examination of other toxin types also revealed similar variability in their presence and activity levels. The high level of venom chemistry variation in varanid lizards compared to that of helodermatid lizards suggests that venom may be subject to different selection pressures in these two families. These results not only contribute to our understanding of venom evolution but also reveal anguimorph lizard venoms to be rich sources of novel bioactive molecules with potential as drug design and development lead compounds.
Asunto(s)
Lagartos , Ponzoñas , Animales , Evolución Molecular , Íleon/efectos de los fármacos , Íleon/fisiología , Técnicas In Vitro , Calicreínas/química , Masculino , Microscopía Electrónica de Rastreo , Contracción Muscular/efectos de los fármacos , Fosfolipasas A2/química , Filogenia , Proteómica , Ratas , Diente/ultraestructura , Ponzoñas/química , Ponzoñas/genética , Ponzoñas/toxicidadRESUMEN
The term "venomics" was coined to describe the global study of venom and venom glands, targeting comprehensive characterization of the whole toxin profile of a venomous animal by means of proteomics, transcriptomics, genomics and bioinformatics studies. This integrative approach is supported by the rapid evolution of protein, RNA and DNA sequencing techniques, as well as databases, knowledge-bases and biocomputing algorithms. The aim of this review is to illustrate advances in the field of venomics during the last decade, addressing each step of the procedure, from sample collection to data treatment. A special focus is made on new perspectives for a better understanding of the venomous function and for fostering the discovery of new venom-derived drug candidates.
Asunto(s)
Genómica , Proteómica , Ponzoñas/genética , Ponzoñas/metabolismo , Animales , Biología Computacional/métodos , Genoma , Genómica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento , Proteómica/métodos , Manejo de Especímenes , Transcriptoma , Ponzoñas/aislamiento & purificación , Ponzoñas/uso terapéuticoAsunto(s)
Antiinfecciosos/química , Antiinfecciosos/farmacología , Péptidos/química , Péptidos/farmacología , Amidas/síntesis química , Amidas/química , Candida/efectos de los fármacos , Membrana Celular/química , Membrana Celular/efectos de los fármacos , Diseño de Fármacos , Escherichia coli/efectos de los fármacos , Pruebas de Sensibilidad Microbiana , Microscopía Inmunoelectrónica , Conformación Molecular , Imitación Molecular , Pliegue de Proteína , Estructura Secundaria de Proteína , Pseudomonas aeruginosa/efectos de los fármacos , Staphylococcus aureus/efectos de los fármacos , Relación Estructura-Actividad , Urea/síntesis química , Urea/químicaRESUMEN
Predatory marine snails of the genus Conus use venom containing a complex mixture of bioactive peptides to subdue their prey. Here we report on a comprehensive analysis of the protein content of injectable venom from Conus consors, an indo-pacific fish-hunting cone snail. By matching MS/MS data against an extensive set of venom gland transcriptomic mRNA sequences, we identified 105 components out of ~400 molecular masses detected in the venom. Among them, we described new conotoxins belonging to the A, M- and O1-superfamilies as well as a novel superfamily of disulphide free conopeptides. A high proportion of the deduced sequences (36%) corresponded to propeptide regions of the A- and M-superfamilies, raising the question of their putative role in injectable venom. Enzymatic digestion of higher molecular mass components allowed the identification of new conkunitzins (~7 kDa) and two proteins in the 25 and 50 kDa molecular mass ranges respectively characterised as actinoporin-like and hyaluronidase-like protein. These results provide the most exhaustive and accurate proteomic overview of an injectable cone snail venom to date, and delineate the major protein families present in the delivered venom. This study demonstrates the feasibility of this analytical approach and paves the way for transcriptomics-assisted strategies in drug discovery.