Advertisement

Neutralizing interaction between heparins and myotoxin II, a lysine 49 phospholipase A2 from Bothrops asper snake venom. Identification of a heparin-binding and cytolytic toxin region by the use of synthetic peptides and molecular modeling.

      This paper is only available as a PDF. To read, Please Download here.
      Heparin binds to phospholipase A2 myotoxins from Bothrops asper snake venom, inhibiting their toxic activities. This interaction was investigated using purified myotoxin II, a Lys-49 phospholipase A2 of this venom, and a series of heparin variants, fragments, and other glycosaminoglycans. The binding was correlated to toxin neutralization, using endothelial cells as a target. Myotoxin II binds radiolabeled heparin in solution unselectively, and forms macromolecular complexes with an optimum at a heparin:toxin molar ratio of 1:5. Both O-sulfates and N-sulfates play a role in heparin binding, in the order of importance 2-O-sulfates > 6-Osulfates > N-sulfates. The shortest heparin oligosaccharides interacting with myotoxin II are hexasaccharides. The binding of a neutralizing monoclonal antibody (MAb-3) to myotoxin II was not inhibited by heparin, indicating that the two molecules interact with different sites on the toxin. A synthetic peptide (residues 115-129 in the numbering system of Renetseder et al. (Renetseder, R., Brunie, S., Dijkstra, B. W., Drenth, J., and Sigler, P. B. (1985) J. Biol. Chem. 260, 11627-11634) of myotoxin II displays both heparin-binding and cytolytic activities. It is concluded that heparin neutralizes myotoxin II by binding to a strongly cationic site in the region of residues 115-129, a possible contribution of lysines 36 and 38 suggested by molecular modeling studies. As this cationic region appears to be responsible for the cytolytic activity of the toxin, the present report constitutes the first identification of a cytotoxic region on a phospholipase A2 myotoxin.

      REFERENCES

        • Rosenberg P.
        Shier W.T. Mebs D. Handbook ofToxinology. Marcel Dekker, Inc., New York1990: 67-277
        • Rosenberg P.
        Harris J.B. Natural Toxins. Clarendon Press, Oxford1986: 129-174
        • Gutierrez J.M.
        • Lomonte B.
        • Cerdas L.
        Toxicon. 1986; 24: 885-894
        • Homsi-Brandeburgo M.I.
        • Queiroz L.S.
        • Santo-Neto H.
        • Rodrigues-Simioni L.
        • Giglio J.R.
        Toxicon. 1988; 26: 615-627
        • Lomonte B.
        • Gutierrez J.M.
        Toxicon. 1989; 27: 725-733
        • Krizaj I.
        • Bieber A.L.
        • Ritonja A.
        • Gubenoek F.
        Eur. J. Biochem. 1991; 202: 1165-1168
        • Kihara H.
        • Uchikawa R.
        • Hattori S.
        • Ohno M.
        Biochem. Int. 1992; 28: 895-903
        • Chwetzoff S.
        • Couderc J.
        • Frachon P.
        • Menez A.
        FEBS Lett. 1989; 248: 1-4
        • Kjell6n L.
        • Lindahl U.
        Annu. Rev. Biochem. 1991; 60: 443-475
        • Lomonte B.
        • Tarkowski A.
        • Bagge U.
        • Hanson L.A.
        Biochem. Pharmacol. 1994; 47: 1509-1518
        • Francis B.
        • Gutierrez J.M.
        • Lomonte B.
        • Kaiser I.I.
        Arch. Biochem. Biophys. 1991; 284: 352-359
        • Lindahl U.
        • Cifonelli J.A.
        • Lindahl B.
        • Roden L.
        J. Biol. Chem. 1965; 240: 2817-2820
        • Hook M.
        • Riesenfeld J.
        • Lindahl U.
        Anal. Biochem. 1982; 119: 236-245
        • Nagasawa K.
        • Inoue Y.
        • Kamata T.
        Carbohydr. Res. 1977; 58: 47-55
        • Lloyd A.G.
        • Embery G.
        • Fowler L.J.
        Biochem. Pharmacol. 1971; 20: 637-648
        • Danishefsky I.
        • Steiner H.
        Biochim. Biophys. Acta. 1965; 101: 37-45
        • Jaseja M.
        • Rej R.N.
        • Sauriol F.
        • Perlin A.S.
        Can. J. Chem. 1989; 67: 1449-1456
        • Inoue Y.
        • Nagasawa K.
        Carbohydr. Res. 1976; 46: 87-95
        • Iverius P.-H.
        Biochem. J. 1971; 124: 677-683
        • Lycke E.
        • Johansson M.
        • Svennerholm B.
        • Lindahl U.
        J. Gen. Virol. 1991; 72: 1131-1137
        • Shively J.E.
        • Conrad H.E.
        Biochemistry. 1976; 15: 3932-3942
        • Pejler G.
        • Lindahl U.
        • Larm O.
        • Scholander E.
        • Sandgren E.
        • Lundblad A.
        J. Biol Chem. 1988; 263: 5197-5201
        • Lane D.A.
        • Denton J.
        • Flynn A.M.
        • Thunberg L.
        • Lindahl U.
        Biochem. J. 1984; 218: 725-732
        • Bitter T.
        • Muir H.M.
        Anal. Biochem. 1962; 4: 330-334
        • Laemmli U.K.
        Nature. 1970; 227: 680-685
        • Reisfeld R.A.
        • Lewis U.J.
        • Williams D.E.
        Nature. 1962; 195: 281-283
        • Valerio R.M.
        • Bray A.M.
        • Campbell R.A.
        • DiPasquale A.
        • Margellis C.
        • Rodda S.J.
        • Geysen H.M.
        • Maeji N.J.
        Int. J. Pept. Protein Res. 1993; 42: 1-9
        • Renetseder R.
        • Brunie S.
        • Dijkstra B.W.
        • Drenth J.
        • Sigler P.B.
        J. Biol. Chem. 1985; 260: 11627-11634
        • Maccarana M.
        • Casu B.
        • Lindahl U.
        J. Biol. Chem. 1993; 268: 23898-23905
        • Bussolino F.
        • De Rossi M.
        • Sica A.
        • Colotta F.
        • Wang J.M.
        • Bocchietto E.
        • Padura I.M.
        • Bosia A.
        • Dejana E.
        • Mantovani A.
        J. Immunol. 1991; 147: 2122-2129
        • Lomonte B.
        • Kahan L.
        Toxicon. 1988; 26: 675-689
        • Lomonte B.
        • Gutierrez J.M.
        • Ramirez M.
        • Diaz C.
        Toxicon. 1992; 30: 239-245
        • Bolanos R.
        • Cerdas L.
        Bol. Oficina Sanit. Panam. 1980; 88: 189-196
        • Holland D.R.
        • Clancy L.L.
        • Muchmore S.W.
        • Ryde T.J.
        • Einspahr H.M.
        • Finzel B.C.
        • Heinrikson R.L.
        • Watenpaugh K.D.
        J. Biol. Chem. 1990; 265: 17649-17656
        • Lindahl U.
        • Thunberg L.
        • Backstrom G.
        • Riesenfeld J.
        • Nordling K.
        • Bjcirk I.
        J. Biol. Chem. 1984; 259: 12368-12376
        • Casu B.
        • Petitou M.
        • Provasoli M.
        • Sina Y.P.
        Trends Biochem. Sci. 1988; 13: 221-225
        • Dua R-
        • Cho W.
        Eur J. Biochem. 1994; 221: 481-490
        • Ibel K.
        • Poland G.A.
        • Baldwin J.P.
        • Pepper D.S.
        • Luscombe M.
        • Holbrook J.J.
        Biochim. Biophys. Acta. 1986; 870: 58-63
        • Maccarana M.
        • Lindahl U.
        Glycobiology. 1993; 3: 271-277
        • Diccianni M.B.
        • Lilly M.
        • McLean L.R.
        • Balasubramaniam A.
        • Harmony J.A.K.
        Biochemistry. 1991; 30: 9090-9097
        • Westerlund B.
        • Nordlund P.
        • Uhlin U.
        • Eaker D.
        • Eklund H.
        FEBS Lett. 1992; 301: 159-164
        • Scott D.L.
        • Achari A.
        • Vidal J.C.
        • Sigler P.B.
        J. Biol. Chem. 1992; 267: 22645-22657
        • Lomonte B.
        • Tarkowski A.
        • Hanson L.A.
        Toxicon. 1994; 32: 1359-1369
        • Bultr6n E.
        • Thelestam M.
        • Gutierrez J.M.
        Biochim. Biophys. Acta. 1993; 1179: 253-259
        • Dijkstra B.W.
        • Kalk K.H.
        • Hoi W.G.
        • Drenth J.
        J. Mol. Biol. 1981; 147: 97-123
        • Scott D.L.
        • White S.P.
        • Otwinowski Z.
        • Yuan W.
        • Gelb M.H.
        • Sigler P.B.
        Science. 1990; 250: 1541-1546
        • Diaz C.
        • Gutierrez J.M.
        • Lomonte B.
        • Gen J.A.
        Biochim. Biophys. Acta. 1991; 1070: 455-460
        • Rufini S.
        • Cesaroni P.
        • Desideri A.
        • Farias R.
        • Gubensek F.
        • Gutierrez J.M.
        • Luly P.
        • Massoud R.
        • Morero R.
        • Pedersen J.Z.
        Biochemistry. 1992; 31: 12424-12430
        • Kaiser T.T.
        • Gutierrez J.M.
        • Plummer D.
        • Aird S.D.
        • Odell G.V.
        Arch. Biochem. Biophys. 1990; 278: 319-325
        • Yoshizumi K.
        • Liu S.Y.
        • Miyata T.
        • Saita S.
        • Ohno M.
        • Iwanaga S.
        • Kihara H.
        Ibxicon. 1990; 28: 43-54
        • Liu S.Y.
        • Yoshizumi K.
        • Oda N.
        • Ohno M.
        • Tokunaga F.
        • Iwanaga S.
        • Kihara H.
        J. Biochem. (Tokyo). 1990; 107: 400-408
        • Maraganore J.M.
        • Merutka G.
        • Cho W.
        • Welches W.
        • K6zdy F.J.
        • Heinrikson R.L.
        J. Biol. Chem. 1984; 259: 13839-13843
        • Kini R.M.
        • Evans H.J.
        Int. J. Pept. Protein Res. 1989; 34: 277-286
        • Kini R.M.
        • Iwanaga S.
        Toxicon. 1986; 24: 895-905