Phospholipase A 2 activity of notexin and its role in muscle damage

Abstract

The phospholipase A 2 activity of notexin, a toxin isolated from the crude venom of Notechis scutatus scutatus, the Australian tiger snake, has been partially characterised using as substrates synthetic dipalmitoyl phosphatidyl choline and bacterial membrane fragments in which the phosphoglycerides were labelled with 1- 14C palmitic acid. The phospholipase activity was Ca 2+-dependent and inhibited by Mg 2+, Ba 2+, Sr 2+, Zn 2+ and Mn 2+; Mg 2+ was the least potent inhibitor and Ba 2+ the most potent. The enzyme was active over a wide range of temperatures and pH, and was stimulated by the presence of deoxycholate. When muscles were incubated in vitro in the presence of notexin, mean muscle fibre potential fell by 16 mV over the first 3 hr and by 60 mV over 6 hr. The depolarisation was Ca 2+-dependent. Muscles were also exposed to notexin in vivo and one day later the phospholipids extracted. Thin layer chromatography revealed the presence of lysophosphatides in the toxin damaged muscles, but not in the contralateral muscles. It was concluded that notexin is a potent phospholipase A 2, capable of hydrolysing phospholipids in micelles, in fragmented bacterial membranes, and in intact muscle fibre membranes, and that this activity represents its primary mode of attack.