Studies on physiological roles of galactolipase in the chloroplasts and cloning of a cDNA encoding galactolipase from Phaseolus vulgaris L.

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In this research project, we studied physiological roles of galactolipase in the stress-induced lipid metabolism, immunological properties of galactolipase, and in addition, we tried to isolate a cDNA encoding galactolipase from kidney bean leaves. The results obtained are as follows :1. The metabolic changes from galactolipids to triacylglycerols are results of the enhancement of galactolipase activity in chloroplasts. The lipid changes was shown to occur not only in spinach but also occurred in leaves subjected to SO_2 treatment, water deficiency, and protoplast isolation.2. At least two types of galactolipase, membrane-bound in chloroplasts and soluble, have been reported in kidney bean and runner bean leaves and prepared rabbit antiserum against this enzyme. The antiserum completely inhibited galactolipase activities not only in the chloroplast fractions but also in the soluble ones. Western blot of both solubilized chloroplast proteins and soluble proteins showed single stained band, suggesting the absence of immunologically different isozymes in both fractions.3. A cDNA library was constructed from poly (A) ^+ RNA which was isolated from younger leaves of kidney bean and purified by oligo (dT)-cellulose column. Early screening of the library using antiserum against galactolipase was unsuccessful. After the solution of the sevsral problems and the reconstruction of the library, we now continue to isolate a cDNA clone encoding galactolipase.