Artigos Publicados em Periódicos

P050-00 Biochemical characterization of two crotamine isoforms isolated by a single step RP-HPLC from Crotalus durissus terrificus (South American rattlesnake) venom and their action on insulin secretion by pancreatic islets Toyama MH, Carneiro EM, Marangoni S*, Barbosa RL, Corso G, Boschero AC* Crotamine. a neurotoxin present in the venom of the South American rattlesnake Crotalus durrisus terrificus exists as several polymorphic variants, as demonstrated by recombinant DNA technology (Smith and Schmidt, Toxicon 28 (1990) 575-585). We have isolated native crotamine by chromatography on Sephadex G75, and have purified two crotamine isoforms by a single step of RP-HPLC. Native crotamine and RP-HPLC fractions F2 and F3 produced skeletal muscle spasms and spastic paralysis in mice. The increase in insulin secretion induced by F2 fraction (5 mu g/ml) was similar to that obtained with 16.5 mu g of native crotamine/ml. These results indicate that the mode of action of the F2 and F3 isoforms in beta-cells is different from that in muscle cells possibly related to the binding affinity of each isoform for the Na+ channels located in the beta-cell membrane. Crotamine isoforms may be valuable tools for studying the involvement of Na+ channels in the mechanism of insulin secretion. Biochimica et Biophysica Acta 1474(1): 56-60, 2000. IF = 2.590 *E-mail: marango@unicamp.br, boschero@unicamp.br

P051-00 Inflammatory oedema induced by the Lys-49 phospholipase A(2) homologue piratoxin-I in the rat and rabbit - Effect of polyanions and p-bromophenacyl bromide Landucci ECT, de Castro RC, Toyama M, Giglio JR, Marangoni S*, De Nucci G, Antunes E Piratoxin-I (PrTX-I) is a Lys-49 phospholipase (PLA(2)) homologue, isolated from Bothrops pirajai snake venom, that has no phospholipase activity. In this study, we investigated the in vivo oedematogenic activity of PrTX-I in both the rat and the rabbit as well as the ability of PrTX-I to activate rat mast cells in vitro. Our results are consistent with the suggestion that the cationic charge of PrTX-I plays a major role in the inflammatory responses induced by this PLA, homologue. Biochemical Pharmacology, 59(10): 1289-1294, 2000. IF = 2.755 *E-mail: marango@unicamp.br

P052-00 Amino acid sequence of piratoxin-II, a myotoxic Lys49 phospholipase A(2) homologue from Bothrops pirajai venom Toyama MH, Soares AM, Wen-Hwa L, Polikarpov I, Giglio JR, Marangoni S* The complete amino acid sequence of the 121 amino acid residues of piratoxin II, a phospholipase A(2) like myotoxin from Bothrops pirajai venom, is reported. PrTX-II is a basic protein with a molecular mass of 13740 Da, a calculated pI of 9.03, but an experimental pI of 8.4 +/- 0.2, showing sequence similarity with other bothropic (90-99%) or non-bothropic (approximate to 80%) Lys49 PLA(2)-like myotoxins. When aligned with B. jararacussu bothropstoxin-I and with B. asper Basp-II, piratoxin-II revealed a single (position 132) and a quintuple (positions 17, 90, 111, 120 and 132) amino acid substitution, respectively, suggesting a common evolutionary origin for these three myotoxins. Biochimie 82(3): 245-250, 2000. IF = 1.594 *E-mail: marango@unicamp.br

P053-00 Leucocyte recruitment induced by type II phospholipases A(2) into the rat pleural cavity de Castro RC, Landucci ECT, Toyama MH, Giglio JR, Marangoni S*, De Nucci G, Antunes E Bothropstoxin-I (BthTX-I) and bothropstoxin-II (BthTX-II) are Lys-49 and Asp-49 phospholipases A(2) (PLA(2)s), respectively, isolated from Bothrops jararacussu venom. Piratoxin-I (PrTX-I) is a Lys-49 PLA? isolated from Bothrops pirajai venom. In this study, the ability of BthTX-I, BthTX-II and PrTX-I to recruit leucocytes into the rat pleural cavity and potential mechanisms underlying this effect were investigated. Our results demonstrate that BthTX-I, BthTX-II and PrTX-I recruit leucocyte into the pleural cavity of the rat by mechanisms unrelated to enzymatic activity and pleural mast cell degranulation. Toxicon 38(12): 1773-1785, 2000. IF = 1.248 *E-mail: marango@unicamp.br

P054-00 Trypsin inhibitor from Dimorphandra mollis seeds: purification and properties Macedo MLR, de Matos DGG, Machado OLT, Marangoni S*, Novello JC* A trypsin inhibitor from Dimorphandra mollis seeds was isolated to apparent homogeneity by a combination of ammonium sulfate precipitation, gel filtration, ion-exchange and affinity chromatographic techniques. SDS-PAGE analysis gave an apparent molecular weight of 20 kDa, and isoelectric focusing analysis demonstrated the presence of three isoforms. The partial N-terminal amino acid sequence of the purified protein showed a high degree of homology with various members of the Kunitz family of inhibitors. This inhibitor is formed by a single polypeptide chain with an arginine residue in the reactive site. Phytochemistry 54(6): 553-558, 2000. IF = 1.106 *E-mail: marango@unicamp.br

P055-00 Effect of a toxic protein isolated from Zea mays seeds on the development and survival of the cowpea weevil, Callosobruchus maculatus Macedo MLR, Coelho MB, Freire MDM, Lima O, Machado T, Marangoni S*, Novello JC* A new toxic glycoprotein (Zeatoxin) has been isolated from Zea mays and sequenced. The toxin is a single polypeptide chain, with Mr of 10 kDa. Zeatoxin was not digested by a mixture of pepsin and papain or by midgut preparations of C. maculatus. A diet of artificial seeds (final concentration, 0.25%) was lethal to 50% of C. maculatus. These results show that Zeatoxin is a new toxic glycoprotein to C. maculatus. A search in the database indicated that the N-terminal sequence show no homology to any other known protein. Protein and Peptide Letters 7(4): 225-231, 2000. IF = 0.557 *E-mail: marango@unicamp.br, jcn@unicamp.br

P056-00 HSP72 as a complementary protection against oxidative stress induced by exercise in the soleus muscle of rats Smolka MB, Zoppi CC, Alves AA, Silveira LR, Marangoni S, Pereira-Da-Silva L, Novello JC, Macedo DV* Given the potential of reactive oxygen species to damage intracellular proteins during subsequent bouts of muscle contractions, it was suggested that, when this production exceeds the antioxidant capacity, the preexisting antioxidant pathways may be complemented by the synthesis of the defense mechanism represented by heat shock proteins (HSPs), stress proteins with the function of repair and maintaining protein folding. To test this hypothesis, reactive carbonyl derivatives were analyzed in plasma and the expression of HSP72 and activities of enzymes from the oxidative and antioxidant defense systems determined in the soleus muscle of sedentary rats and rats trained by two protocols: continuous and intermittent. We analyzed all three groups at rest and 2 h after acute exercise. After 8 wk of training, the animals from both groups clearly demonstrated higher resistance to exercise. Both trained groups showed significantly higher citrate synthase, catalase, and glutathione reductase activities than the control group (P < 0.01). After acute exercise, the induction of HSP72 expression occurred only in the sedentary group, suggesting that HSP72 acts as a complementary protective mechanism in exercise-induced oxidative stress. American Journal of Physiology-Regulatory Integrative and Comparative Physiology 279(5): 1539-1545, 2000. IF = 2.453 *E-mail: labex@unicamp.br

P057-00 Isolation and characterization of novel neurotoxins from south American rattlesnake Crotalus durissus terrificus: Optimization of chromatographic procedure Toyama MH, Leite GB, Rodrigues-Simioni L, Hernandez OSS, Novello JC*, Marangoni S* Molecular exclusion HPLC chromatography on a Protein Pak SW 300 column and reversed phase HPLC on a mu-Bondapack C18 column were used to purify four novel neurotoxins from the crotoxin complex, and five crotoxin subunits (PLA(2) and crotapotin) as well as two crotamine isoforms. A single preparative reversed phase HPLC steps yielded a major basic myotoxin followed by the subunits of crotoxin (three PLA(2) isoforms and two crotapotin isoforms). This modified procedure may be useful for rapid, large-scale purification of toxins from Crotalid venoms. Protein and Peptide Letters 7(6): 381-388, 2000. IF = 0.557 *E-mail: marango@unicamp.br; jcn@unicamp.br

P058-00 Radicais livres de oxigênio: um software introdutório Yokaichiya DK, Galembeck E*, Torres BB Química Nova 23(2): 267-269, 2000. IF = 0.304 *E-mail: edgalemb@unicamp.br

P059-00 Tetracaine stimulates insulin secretion through the mobilization of Ca2+ from thapsigargin- and IP3- insentitive Ca2+ rservoir in pancreatic b-cells Bosqueiro, JR, Carneiro EM, Bordin S, Boschero AC* The effect of tetracaine on 45Ca fluxes, cytoplasmic Ca2+ concentrations [Ca2+]i and insulin secretion in isolated pancreatic islets and B-cells was studied. Our data indicate that tetracaine mobilizes Ca2+ from a tapsigargin-insensitive store and hence stimulates insulin secretion even in the absence of Ca2+. The increase in 45Ca efflux provoked by high K+ and by Cch indicates that tetracaine did not interfere neither with a cation nor with an IP3-sensitive Ca2+ pool in B-cells. Canadian Journal of Physiology and Pharmacology 78: 462-468, 2000. IF= 1.493 *E mail: boschero@unicamp.br

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