Cloning Of A Novel ω-conotoxin Gene MiEN

Conotoxins (CTXs) are small, structured peptide toxins, which are generated by the predatory cone snails (genus Conus) for prey capture, defense, and competitor deterrence. Venomous Conus are tropical marine mollusks, and native to Hainan only in China. Conopeptides (conotoxins) are widely used as research tools in neuroscience, and used as new potential therapeutic agents, because of their structure novelty, strong bioactivities and unprecedented selectivity in targeting specific molecular forms of either voltage-gated or ligand-gated ion channels (and in a few cases, G-protein linked receptors). Vermivorous Conus toxin libraries represent a rich insecticidal mine. Conotoxins are the potential to "therapeutically" such a resource appears limitless.Conotoxins can be grouped into A-, O-, T-, M-, P-, I- superfamilies et al, according to highly conserved signal peptide of conotoxin precurser, which are also can be classified into α-, ω-, μ- and δ- family subtypes etc. according to different targets they block . co-conotoxins can block and discriminate a diverse set of different molecular isoforms of voltage-gated calcium channels, which are widely used as pharmacological tools in neuroscience for relative ion channel researches, some has been through extensive human clinical trials. Researches on co-conotoxins (ω-CTX) flourish now, which become the marine drugs' research foucus. The cysteine framework of co-conotoxins is -C-C-CC-C-C-, composed of 24-31 amino acids, each containing three pairs of disulfide bonds which constitute a four-loop framework. Researches on insecticidal resistance activity of omega-conotoxins have never been reported up to now.In this research different methods are adopted to exstract DNA and RNA to set up a set of optimizing plan suitable to the extraction of conotoxin nucleic acid. A novel co -conotoxin conotoxin cDNA (MiEN gene) was isolated from venom mRNA of Conus miles Linnaeus native to Hainan by 3' -RACE. Amino acid sequence of mature peptide MiEM was deduced from MiEN cDNA. The synthesis of the linear peptide MiEM was performed using the Fmoc strategy. Bioactivities of MiEM to cockroach (Periplaneta Americana) were assayed after mature peptide folding correctly. The results showed that MiEM had lethal effect on cockroach. The LD50 (i.e. the dose that is lethal to 50% of...