Yon-Hui Lin

Lipase in lipid bodies of cotyledons of rape and mustard seedlings

Lipolytic activity was absent in the crude cotyledon extract of ungerminated rapeseed (Brassica napus L. var. Dwarf Essex), and increased to a peak at day 4 in seedling growth, concomitant with the decrease in total lipids. About 50% of the lipase activity was recovered in the lipid bodies isolated from the cotyledon extract by flotation centrifugation. Isolated lipid bodies underwent autolysis of internal triacylglycerols resulting in the release of fatty acids. After the triacylglycerols in isolated lipid bodies had been extracted with diethyl ether, the lipase was recovered in the remaining membrane fraction. The lipase had a maximal activity at pH 6.5 on trierucin, trilinolein, or endogenous triacylglycerols, and at pH 8.0 on N-methylindoxylmyristate. The lipase was most active on trierucin and trilinolein, and hydrolyzed the related di- and monoacylglycerols at lower rates. There was little enhancement of the lipase activity in the presence of NaCl, CaCl2, or detergents, and detergents in general reduced the activity. The hydrolysis of trierucin was linear until about 50% of the trierucin had been converted to erucic acid, and there was little accumulation of dierucin and monoerucin. Lipase extracted from lipid bodies isolated from germinated rapeseed of the variety Tower, which contains little or no erucic acids in the storage triacylglycerols, also had the highest activities on trierucin and trilinolein. A comparative study on mustard seed (Brassica juncea) revealed that the mustard lipase possessed characteristics very similar to those of the rapeseed lipase.

Characteristics and biosynthesis of seed lipases in maize and other plant species

Oilseed lipases from diverse plant species exhibit differences in their substrate specificity, pH for optimal activity, reactivity toward sulfhydryl reagents, hydrophobicity and subcellular location. Seed lipase from a certain plant species is relatively specific for the native triacylglycerols or triacylglycerols containing the major fatty acids of the storage triacylglycerols of the same species. This substrate specificity can be exploited in lipid biotechnology. In most seeds, with the known exception of castor bean, lipase activities are absent in ungerminated seeds and increase in postgermination. The biosynthesis of seed lipase has been studied only in maize. The maize enzyme is synthesized on free polyribosomes in postgermination. The newly synthesized enzyme is then transferred, without apparent coor posttranslational modification, to the membrane of the lipid bodies.

Synthesis and Degradation of Lipid Bodies in the Scutella of Maize

Most seeds contain storage lipids in the form of triacylglycerols, which usually comprise 20–50% of the total seed dry weight (1, 2, 3, 4). This lipid reserve is synthesized during seed maturation, and is rapidly mobilized to provide energy and carbon skeleton for the growth of the embryo during germination. The triacylglycerols are densely packed in subcellular organelles called lipid bodies (oleosomes, spherosomes, oil bodies). The spherical lipid body is about 0.5–1 μm in diameter, and is surrounded by a “half-unit” membrane of one monolayer of phospholipids about 3 nm thickness (5). The fatty acyl moieties of the membrane phospholipids are believed to orient themselves toward the matrix so that they can form hydrophobic interaction with the internal triacylglycerols.

Substrate Specificity of Plant Lipases

In plants, lipases (EC 3.1.1.3) are active in lipid-storing tissues in seeds during germination. These lipases catalyze the hydrolysis of storage triacylglycerols to fatty acids, which are converted to sugars to support the growth of the embryo. Seed lipases have not been studied intensively, and our knowledge of them lags behind that of the mammalian and microbial enzymes (1, 2, 3). A unique feature of seed lipases may be that the enzyme from a certain plant species is relatively specific on the characteristic storage triacylglycerols of the same species. Our experimental results show that this specificity is indeed species-dependent, and is not due solely to the physical characteristics of the specific triacylglycerol in the substrate micelles.