Biochemical and biological properties of phospholipases A(2) from Bothrops atrox snake venom.
Phospholipases A(2) (PLA(2)s), of molecular mass 13-15kDa, are commonly isolated from snake venom. Two myotoxins with PLA(2) activity, BaPLA(2)I and BaPLA(2)III, with estimated molecular masses of 15kDa were isolated from the venom of Bothrops atrox using Sephacryl S-100-HR and reverse-phase chromatography. BaPLA(2)I was basic, with a pI of 9.1, while BaPLA(2)III was neutral with a pI of 6.9. On a molecular basis, BaPLA(2)III exhibited higher catalytic activity on synthetic substrates than BaPLA(2)I. Comparison of the N-terminal residues of BaPLA(2)I with other PLA(2) proteins from snake venoms showed that it has the highest homology (94%) with B. asper myotoxin II and homology with a PLA(2) Lys(49) from B. atrox (89%). In contrast, BaPLA(2)III demonstrated 75, 72, and 71% homology with PLA(2) from Vipera ammodytes meridionalis, B. jararacussu, and B. jararaca, respectively. BaPLA(2)I and BaPLA(2)III were capable, in vitro, of inducing mast cell degranulation and, in vivo, of causing creatine kinase release, edema, and myonecrosis typical of PLA(2)s from snake venoms, characterized by rapid disruption of the plasma membrane as indicated by clumping of myofilaments and necrosis of affected skeletal muscle cells. BaPLA(2)I- and BaPLA(2)III-specific monoclonal and polyclonal antibodies, although incapable of neutralizing PLA(2) edematogenic activity, blocked myonecrosis efficiently in an in vivo neutralization assay. The results presented herein suggest that the biological active site responsible for edema induction by these two PLA(2) enzymes is distinct from the myonecrosis active site and is not dependent upon the catalytic activity of the PLA(2) enzyme.
Kanashiro
Milton M
MM
Laboratório de Biologia do Reconhecer, Centro de Biociências e Biotecnologia - LBR, Universidade Estadual do Norte Fluminense, Av. Alberto Lamego, 2000, Campos dos Goytacazes, Rio de Janeiro, Brazil.
de Cássia M Escocard
Rita
R
Petretski
Jorge H
JH
Prates
Maura V
MV
Alves
Elias W
EW
Machado
Olga L T
OL
da Silva
Wilmar Dias
WD
Kipnis
Thereza L
TL
D000595:Amino Acid Sequence; D000818:Animals; D017837:Bothrops; D002384:Catalysis; D003402:Creatine Kinase / Q000378:metabolism; D003435:Crotalid Venoms / Q000201:enzymology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009130:Muscle, Smooth / Q000187:drug effects / Q000473:pathology; D010741:Phospholipases A / Q000737:chemistry / Q000378:metabolism*; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid
D003435:Crotalid Venoms; D003402:Creatine Kinase; D010741:Phospholipases A