Characterisation of neurotoxic polypeptides from Cyanea capillata medusae (Scyphozoa)

Abstract

Cnidarian venoms include neurotoxins, which are able to paralyse prey organisms immediately. Important targets for neurotoxins are voltagegated ion channels in membranes of excitable cells. By blocking specific receptor sites, neurotoxic components disturb the physiological ion channel functions. Here, we describe the isolation and characterisation of potential neurotoxic polypeptides from the crude tentacle venom of the boreal scyphomedusan Cyanea capillata. Partially purified venom fractions were obtained by size-exclusion and subsequent reversedphase chromatography. To assess the blocking activity of the venom on voltage-gated sodium channels, we modified a mouse neuroblastoma (MNB) cell assay. Venom fractions containing channel-blocking activity were analysed by matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS). The resulting mass spectra revealed a cluster of singly charged peptides within a mass range from 3,900 to 7,000 Da. A group of three potentially neurotoxic peptides with molecular masses of 3983.4, 5795.4 and 6962.1 Da could be tracked throughout the purification process. This investigation of the crude venom is part of a multidimensional assay-guided approach for the isolation and structural characterisation of toxic polypeptides in northern Scyphozoa.

Gill cell toxicity of northern boreal scyphomedusae Cyanea capillata and Aurelia aurita measured by an in vitro cell assay

“…This may be related to the presence of proteolytic enzymes in the oral arms, which function as digestive enzymes and affect the gill cells nonspecifically, whereas the toxins in the fishing tentacles include various neurotoxins to induce rapid paralysis of prey organisms which probably do not affect the gill cells in the same way (see Lassen et al, 2010).…”

Isolation of a Nav channel blocking polypeptide from Cyanea capillata medusae – A neurotoxin contained in fishing tentacle isorhizas

“…The preparation of the crude venom from thawed nematocyst suspensions of WI, its initial fractionation by size-exclusion chromatography (SEC), the concentration and desalting of SEC fractions, as well as the first purification step by reversed-phase chromatography (RPC1) were carried out according to Lassen et al (2010). RPC1 fractions of several LC runs (injection volume 100 μL) were combined correspondingly, concentrated and investigated in terms of neurotoxic activity (see 2.3) or stored at -28 °C for further purification.…”

“…The released MTT derivative was immediately measured in a multiwell scanning photometer (Victor 3, Perkin Elmer, Germany) at 550 nm. The cell assay principle and the calculation of the neurotoxic activity (NA) are described in Lassen et al (2010). In brief, the 6 NA of each fraction and concentration, respectively, was calculated based on the median absorbance values of the eight replicates per test in the following way:…”

“…A first investigation of C. capillata medusae regarding their neurotoxic activity was carried out by Lassen et al (2010). They showed a blocking activity on voltage-gated sodium (Na v ) channels expressed by mouse neuronal cells in partially purified fishing tentacle venom.…”

Biochemical and Electrophysiological Characterization of Two Sea Anemone Type 1 Potassium Toxins from a Geographically Distant Population of Bunodosoma caissarum

“…As the most ancient venomous animals on Earth, cnidarians (classes Anthozoa, Scyphozoa, Cubozoa and Hydrozoa) have evolved a large amount of pore-forming toxins, phospholipases A 2 , protease inhibitors, neurotoxins and toxic secondary metabolites [1,2,3,4,5,6]. The biological and ecological roles of these toxins present in cnidarians venom are (1) immobilization and death of the prey, (2) defense against predators and (3) intra- and inter-specific competition [7,8,9,10,11].…”