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Title: Two novel dermonecrotic toxins LiRecDT4 and LiRecDT5 from Brown spider (Loxosceles intermedia) venom: From cloning to functional characterization
Authors: Silveira, Rafael Bertoni da
Pigozzo, Romine Bachmann
Chaim, Olga Meiri
Appel, Marcia Helena
Silva, Dilza Trevisan
Dreyfuss, Juliana Luporini
Toma, Leny
Dietrich, Carl Peter
Nader, Helena B.
Veiga, Silvio Sanches
Gremski, Waldemiro
Univ Fed Parana
Universidade Federal de São Paulo (UNIFESP)
Catholic Univ Parana
Keywords: Brown spider
Loxosceles intermedia
dermonecrotic toxin
phospholipase activity
Issue Date: 1-Mar-2007
Publisher: Elsevier B.V.
Citation: Biochimie. Paris: Elsevier France-editions Scientifiques Medicales Elsevier, v. 89, n. 3, p. 289-300, 2007.
Abstract: Loxoscelism (the condition produced by the bite of brown spiders) has been reported worldwide, but especially in warmer regions. Clinical manifestations include skin necrosis with gravitational spreading while systemic loxoscelism may include renal failure, hemolysis and thrombocytopenia. the venom contains several toxins, of which the best biochemically and biologically studied is the dermonecrotic toxin, a phospholipase-D. Purified toxin induces cutaneous and systemic loxoscelism, especially necrotic lesions, hematological disturbances and renal failure. Herein, we describe cloning, heterologous expression and purification of two novel dermonecrotic toxins: LiRecDT4 and LiRecDT5. the recombinant proteins stably expressed in Escherichia coli cells were purified from culture supernatants in a single step using Ni(-)(2+)chelating chromatography producing soluble proteins of 34 kDa (LiRecDT4) and 37 kDa (LiRecDT5). Circular dichroism analysis evidenced correctly folding for toxins but differences in secondary structures. Both proteins were recognized by whole venom serum antibodies and by a specific antibody to dermonecrotic toxin. Also, recombinant toxins with phospholipase activity induced experimental skin lesions and caused a massive inflammatory response in rabbit skin dermis. Nevertheless, toxins displayed different effects upon platelet aggregation, increase in vascular permeability and not caused death in mice. These characteristics in combination with functional studies illustrates that a family of dermonecrotic toxins exists, and includes two novel members that are useful for future structural and functional studies. They will also be useful in biotechnological ends, for example, as inflammatory and platelet aggregating studies, as antigens for serum therapy source and for lipids biochemical research. (c) 2007 Elsevier Masson SAS. All rights reserved.
ISSN: 0300-9084
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