Biopolym. Cell. 2005; 21(4):319-325.
Study on carbohydrate specificity of hemolytic lectin from death-cap mushroom (Amanita phalloides (Vaill. Fr.) Secr)
- Institute of Cell Biology, NAS of Ukraine
14/16, Drahomanov Str., Lviv, Ukraine, 79005
Purification and some physical and chemical properties of a hemolytic lectin from death-cap mushroom (A. phalloides (Vaill. Fr.) Secr) are described. According to the SDS-PAGE data, the polypeptide chain of phallolysin consists of one component with molecular weight of approximately 18 kDa. The molecular weight of the lectin as determined by gel filtration on Toyopearl HW-55 is 35 kDa. Phallolysin is thermolabile, the heating during 30 min at 65 °C removes completely its hemolytic and hemaglutinating activity. Phallolysin is not Ca2+-dependent lectin. The sensitivity of erythrocyte of different species to the hemolysis decreases in the following sequence: rabbit>rat>man. Osmotic protection experiments were performed using rabbit erythrocytes. When the erythrocytes were incubated together with phallolysin in the presence of polyethylene glycols of different molecular weight, the lysis was inhibited increasingly as the size of the molecules increased. The obtained results indicate that phallolysin forms ion-permeable pores with a functional diameter smaller than 2.3 nm (but big ger than 1.6 nm) in the cell membranes of rabbit erythrocytes. At the same time the presence of polyethylene glycols does not influence the hemaglutinating activity that allows to study carbohydrate specificity of the death-cap mushroom lectin. The lectin interaction with D-galactose and its β-derivatives is the strongest as revealed by study on carbohydrate specificity of phallolysin. The data on the phallolysin interaction with glycoproteins do not show any preference towards either N- or O-glycan type.
Keywords: Amanita phalloides, phallolysin, hemotys, osmotic protect, carbohydrate specificity
 Vasser SP. Flora mushrooms Ukraine. Amanita mushrooms. Kiev: Naukova Dumka, 1992; 168 p.
 Seeger R. Demonstration and isolation of phallolysin, a haemolytic toxin from Amanita phalloides. Naunyn Schmiedebergs Arch Pharmacol. 1975;287(3):277-87.
 Lutsik-Kordovsky MD, Antonyuk VA, Stasyk TV, Yakymovych MYa, Yakymovych LA, Hellman U, Soushelriytsky S, Stoika RS, Lutsik AD. Hemagglutinating lectin from fruit bodies of Amanita phalloides: isolation, properties and, biological activity: Abstr. Portsmouth: Univ. press. 1999: 77.
 Lutsik-Kordovsky MD, Stasyk TV, Stoika RS. Analysis of cytotoxicity of lectins and non-lectin proteins from Amanita mushrooms. Exp Onkol. 2001; 23(1): 43-5.
 Hatakeyama T, Kohzaki H, Nagatomo H, Yamasaki N. Purification and characterization of four Ca(2+)-dependent lectins from the marine invertebrate, Cucumaria echinata. J Biochem. 1994;116(1):209-14.
 Roch P, Canicatti C, Valembois P. Interactions between earthworm hemolysins and sheep red blood cell membranes. Biochim Biophys Acta. 1989;983(2):193-8.
 Tateno H, Goldstein IJ. Molecular cloning, expression, and characterization of novel hemolytic lectins from the mushroom Laetiporus sulphureus, which show homology to bacterial toxins. J Biol Chem. 2003;278(42):40455-63.
 Kouriki-Nagatomo H, Hatakeyama T, Jelokhani-Niaraki M, Kondo M, Ehara T, Yamasaki N. Molecular mechanism for pore-formation in lipid membranes by the hemolytic lectin CEL-III from marine invertebrate Cucumaria echinata. Biosci Biotechnol Biochem. 1999;63(7):1279-84.
 As. SSSR 1554961. A method of producing an affinity sorbent for lectins purification. VA Antonyuk. BI. 13.
 Maurer H. Disc electrophoresis and related techniques of polyacrylamide gel electrophoresis. de Gruyter. BerlinвЂ“New York. 1971. 222 p.
 Kabat EA, Mayer MM. Experimental Immunochemistry. Illinois: Charles Thomas Publ., 1964. 690 p.
 Lazur'evskiy GV, Terent'eva IV, Shamchurin AA. Practical work on Natural Compounds Chemistry. M.: Vyshchaya Shkola. 1966; 336 p.
 Methods of carbohydrate chemistry. Ed. NK Kochetkov. M. Mir, 1967. 512 p.
 Lutsik AD, Detyuk AS, Lutsik MD. Lectin in histochemistry. Lviv: Lviv Univ. publ., 1989 144 p.
 Scherrer R, Gerhardt P. Molecular sieving by the Bacillus megaterium cell wall and protoplast. J Bacteriol. 1971;107(3):718-35.
 Nagahama M, Hayashi S, Morimitsu S, Sakurai J. Biological activities and pore formation of Clostridium perfringens beta toxin in HL 60 cells. J Biol Chem. 2003;278(38):36934-41.