Abstract
Cytotoxic protein from fruit body of the death cap Amanita phalloides mushroom, designated as toxophallin, has been isolated and its principal physico-chemical and biological properties have been characterized. This protein possesses molecular mass 55 kDa and isoelectric point at pH 5.7 and is expressed predominantly in the stem of mushroom fruit body. As assessed by cross-linking experiments, toxophallin forms a trimeric complex in buffered solution. It induces apoptosis in several mammalian cells lines (murine L1210 and CCL-64, and human A549) with IC 50 at 0.25–0.45 ug/ml. It also induces DNA fragmentation and morphological changes in the nuclei of target cells (chromatin condensation and fragmentation of nuclei) that are characteristic for apoptosis. Caspase III Inhibitor (Boc-D-fmk) does not inhibit toxophallin-induced apoptotic DNA fragmentation, suggesting that this toxin involves caspase-independent pathway of apoptosis. Besides, it was shown that toxophallin interaction with target cells is not mediated by specific cell surface receptor. Thus, toxophallin is a new toxic protein whose properties distinguish it from other toxic compounds (cyclopeptides and phallolysin) earlier found in the death cap. Keywords: toxic protein, death cap Amanita phalloides , apoptosis.
Highlights
Death cap (Amanita phalloides L.) is known as a deadly poisonous mushroom due to the presence of several toxic substances
Wieland Th. revealed other toxic cyclopeptides which were classified into two structural groups – amanitine and phalloidine, exhibiting different mechanisms of toxic action [21]
For elimination of cytolytic lectin, phallolysin, the crude protein fraction was passed through a column filled with affinity sorbent – ovomucin immobilized on agarose [2], that was equilibrated with PBS
Summary
Death cap (Amanita phalloides L.) is known as a deadly poisonous mushroom due to the presence of several toxic substances. While amanitin is blocking mRNA transcription, phalloidin binds to actin and alters cytoskeleton functions These cyclopeptides are frequently used as tools in scientific studies since their intracellular molecular targets and mechanisms of action are well characterized. It should be noted that besides toxic peptides this mushroom contains antitoxin antamanide, a cyclodecapeptide which blocks phalloidin effects [1, 21]. Later on, another toxic polypeptide substance, phallolysin, possessing hemolytic activity was detected in the fruit bodies of the death cap [4, 5, 16,17,18]. Its chemical properties and biological activity, as well as the mechanism of action, are well studied [8, 21]
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