163 Biochimica et Biophysica Acta, 512 (1978) 163--171 © Elsevier/North-Holland Biomedical Press BBA 78116 THE SYNERGISM OF CARDIOTOXIN AND PHOSPHOLIPASE A2 IN HEMOLYSIS ABRAHAM I. LOUW and LEON VISSER Molecular Biochemistry Division, National Chemical Research Laboratory, Council for Scientific and i'ndustrialResearch, P.O. Box 395, Pretoria 0001 (Republic of South Africa) (Received November 10th, 1977) (Revised manuscript received January 31st, 1978) Summary The synergistic effect of exogenous cobra phospholipase A2 on the hemol- ysis rate of guinea pig erythrocytes by highly purified snake venom cardio- toxins was investigated. In the presence of phospholipase A2 the reaction was not only faster and had a lower activation energy but followed a sigmoidal instead of a linear time course. Similar results were obtained using porcine pan- creatic phospholipase A2. Significantly, addition of even a trace of cobra phos- pholipase A2 (approx. 0.1%, w/w) was sufficient to bring about the full syner- gistic effect, emphasizing the stringent purity requirements for any meaningful investigation of cardiotoxin's own action. The possibility that the action of cardiotoxin on its own may involve the stimulation of an endogenous phos- pholipase is discussed in the light of the results obtained with exogenous cobra enzyme. Introduction Cardiotoxins represent the principal protein component of cobra venoms (up to 50%, on a weight basis}. The toxicity of this group of low molecular weight proteins is closely related to the marked permeability changes that they induce in the membranes of a variety of cell types [1--6], which in the case of heart muscle result in cardiac arrest. It is also well documented that phospholipases A2 can act synergistically with cardiotoxins to enhance these effects, e.g. in the hemolysis of erythrocytes [7,8] and the depolarization of skeletal muscle [9]. These observations raised several questions about the mechanism by which phospholipases exert a synergistic effect on cardiotoxin action on the one hand and about the possibility, on the other hand, that cardiotoxins may stimulate an endogenous membrane phospholipase to facilitate their in vivo action.