- Citado por SciELO
versão impressa ISSN 0104-7930versão On-line ISSN 1678-4936
J. Venom. Anim. Toxins v. 3 n. 2 Botucatu 1997
Mini-Symposium - Abstracts
2 COMPLETE AMINO ACID SEQUENCE OF AN ASP49 PHOSPHOLIPASE A2 FROM Bothrops erythromelas VENOM.
J. PRADO-FRANCESCHI , C.A. FLORES , S.E. MORENO , J.C. COGO , S. HYSLOP , J.R. GIGLIO , C. BLOCH JR. , L. MORHY
Department of Pharmacology, University of Campinas, Campinas, SP; Department of Biochemistry, University of São Paulo, Ribeirão Preto, SP and Brazilian Center of Protein Sequencing, University of Brasília, Brasília, DF, Brazil
Most phospholipases A2 (PLA2) have the amino acid aspartic acid (Asp) at position 49 in their sequence, which appears to be important for the maintenance of full enzymatic activity. In recent years, an increasing number of PLA2s with lysine (Lys) at position 49 have been identified, and in these enzymes, the level of activity is greatly reduced or apparently absent. In the present work, we have determined the complete amino acid sequence of an Asp49 phospholipase A2 homologue from the venom of Bothrops erythromelas (jararaca da serra). Fractionation of B.erythromelas venom (250 mg) on Sephadex G-75 using ammonium bicarbonate buffer (0.1M, pH 8.0) yielded six principal peaks, of which the third and fourth peaks possessed phospholipase activity and induced neutrophil migration in rats. Alkylation of material from the fourth peak with 4-vinylpyridine followed by reversed phase HPLC on a Vydac C4 column resulted in five peaks of which the fifth peak, which was also the largest one corresponding to 27% of the material loaded onto the column, was selected for sequencing in an automated protein sequencer. N-Terminal amino acid determination revealed the presence of only one amino acid (Ser) indicating that this peak consisted of only one protein. Amino acid analysis demonstrated that the protein contained 121 amino acids with a calculated molecular weight of 13,655. Complete sequencing showed the presence of Asp at position 49, as well as the existence of Cys residues at positions commonly found in Asp49 PLA2. There was a 65% homology between the sequence of the B. erythromelas protein and a homologous enzyme from B. asper. The level of PLA2 activity of this protein remains to be determined as does its involvement in the in vivo neutrophil migration seen in fraction 4 following chromatography of the venom on Sephadex G-75.
Dra. Júlia Prado Franceschi - UNICAMP, Faculdade de Ciências Médicas, Departamento de Farmacologia, Caixa Postal 6111, CEP 13084-100, Campinas, SP, Brasil. email: firstname.lastname@example.org