Rapeseed lipase-pH dependent specificity for native lipid body autolysis and lipolysis of artificial oil droplets

In homogenates of cotyledons from 4-day-old rapeseed (Brassica napus L.) seedlings lipase (EC 3.1.1.3) activity is present in both the soluble cell fraction (recovery about 70%) and the oil body fraction. In contrast to lipases and esterases of mammalian or microbial origin lipase activity from cotyledons of rapeseed seedlings is not inhibited by serine hydrolase inhibitors. Optimum activity with a triolein emulsion as substrate was found at pH 9.0. Addition of desoxycholate to the triolein substrate emulsion, which improves the stabilization of the surface of the lipid globules, shifts the apparent pH optimum to pH 8.0. In contrast, rapeseed oil emulsified in gum arabic was a poor substrate for the soluble lipase. Only after addition of desoxycholate to the emulsified rapeseed oil was the apparent activity of lipase clearly enhanced, and fatty acids, 1,2(2,3)-diacylglycerols, and monoacylglycerols were produced at such alkaline pH values. The autolysis of a crude oil body fraction demonstrating the lipolytic activity associated with these subcellular compartments by the liberation of free fatty acids from the native neutral lipids with maximum activity at pH 5.0 is taken as evidence for the presence of lipase activity in or on the surface of the oil bodies. When assayed with the standard pH-stat method the oil body fraction showed about 8% of the total lipase activity of the crude homogenate, and the enzyme activity was found to be bound only loosely to the surface of the oil bodies. Possible misinterpretations in earlier reports about both the <microsomal localization> and properties of the rapeseed lipase are discussed.