Cyto-insectotoxins, a novel class of cytolytic and insecticidal peptides from spider venom

Abstract

Eight linear cationic peptides with cytolytic and insecticidal activity, designated cyto-insectotoxins (CITs), were identified in Lachesana tarabaevi spider venom. The peptides showed antibiotic activity towards Gram-positive and Gram-negative bacteria at micromolar concentrations as well as toxicity to insects. The primary structures of the toxins were established by direct Edman sequencing in combination with enzymatic and chemical polypeptide degradation and MS. CITs represent a novel class of cytolytic molecules and spider venom toxins. They are the first example of molecules showing equally potent antimicrobial and insecticidal effects. Analysis of L. tarabaevi venom gland expressed sequence tag database revealed the primary structures of the protein precursors; eight peptides homologous with the purified toxins were additionally predicted. CIT precursors share a conventional prepropeptide structure with an acidic prosequence and a processing motif common to most spider toxin precursors. The most abundant peptide, CIT 1a, was chemically synthesized, and its lytic activity on different bacterial strains, human erythrocytes and lymphocytes, insect cells, planar lipid bilayers and lipid vesicles was characterized. The spider L. tarabaevi is suggested to have evolved to rely on a unique set of linear cytolytic toxins, as opposed to the more common disulfide-containing spider neurotoxins.

Spider toxins comprising disulfide‐rich and linear amphipathic domains: a new class of molecules identified in the lynx spider Oxyopes takobius

“…Samples were analyzed by MALDI-MS using M@LDI-LR (Micromass, Manchester, UK) and Ultraflex TOF-TOF (Bruker Daltonik GmbH, Bremen, Germany) spectrometers as described previously [18].…”

“…This signature may be reduced to the more general principal structural motif CX 6 CX n CC (C 1 X 6 C 2 X 5 C 4 C 5 in our case), and extra structural motif CXCX n CXC (C 6 XC 7 X m C 8 XC 9 in our case) that are characteristic of spider neurotoxins [23]. The principal structural motif and the extra structural motif in turn conform to the more general ICK motif CX 2-7 CX 3-11 CX 0-7 CX 1-17 CX [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] …”

“…In addition, there are two possible 'homo' variants: AMP+AMP and ICK+ICK. The first identified examples of modular spider toxins were cyto-insectotoxins in the venom of L. tarabaevi, which contain two linear modules (AMP+AMP) [18]. Subsequently, two-domain spider toxins containing two knottin modules (ICK+ICK) were discovered in the venom of Cheiracanthium punctorium (Miturgidae) and Haplopelma schmidti (Theraphosidae) [16,19].…”

“…CD spectra were obtained on a J-810 spectropolarimeter (Jasco, Tokyo, Japan) as described previously [18].…”

“…We are particularly interested in the variety of so-called 'modular' toxins that comprise two modules or domains, each corresponding to a 'usual' spider toxin. To date, we have described the following combinations: (a) ICK+ICK, two ICK domains in one toxin [16], (b) ICK+AMP, an ICK domain with a long linear C-terminal extension conferring membrane activity [17], and (c) AMP+AMP, two short linear MAPs joined into one long structure [18]. Other research groups have also described modular toxins of the ICK+ICK [19] and ICK+AMP types [20].…”