Biosynthesis of Nitrogenous Phospholipids in Spinach Leaves

Abstract

Pathways for biosynthesis of phosphatidylserine (PS), phosphatidylethanolamine (PE), and phosphatidylcholine (PC), in spinach leaves have been studied both in vivo (whole leaves and leaf slices) and in vitro (cell-free leaf fractions). Biosynthesis of PS was shown to occur by the action of a particle-bound CDP-diglyceride: serine phosphatidyltransferase, and PE by the action of a PS-decarboxylase localized in the 100 000 × g supernatant fraction. PE was also formed by the operation of the CDP-ethanolamine:diglyceride phosphorylethanolamine transferase, localized in the microsomal fraction. The presence of ethanolamine kinase required for formation of phosphorylethanolamine was demonstrated in vitro, but not the presence of CTP:phosphorylethanolamine cytidyltransferase; however, the latter is presumed present on the basis of in vivo results. Operation of the methylation pathway for biosynthesis of PC was established in vivo, and direct methylation of phosphatidyl-N-methylethanolamine to phosphatidyl-N,N-dimethylethanolamine (PE-diMe) and of PE-diME to PC by S-adenosylmethionine was demonstrated with a particulate enzyme system localized in the microsomal fraction; direct methylation of PE itself could not be shown in this system. PC was also synthesized by the CDP-choline:diglyceride phosphorylcholine transferase system localized in the microsomal fraction. Synthesis of PE and PC by Ca2+-stimulated exchange reactions with ethanolamine and choline, respectively, could be demonstrated, but at low rates. However, no synthesis of PS by exchange reactions with serine could be detected.

Synthesis of Ethanolamine and Its Regulation in Lemna paucicostata

“…PtdSerine decarboxylase activity has been demonstrated in extracts of spinach leaves, chiefly in the 100,000g supernatant fraction (26,27 Plant Physiol. Vol.…”

“…PtdSerine decarboxylase activity has been demonstrated in extracts of spinach leaves, chiefly in the 100,000g supernatant fraction (26,27). When coupled with a base-exchange reaction (an activity demonstrated in endoplasmic reticulum from castor bean [28]): serine + PtdEA -* PtdSerine + EA (reaction 2) a cycle can be established, the net effect being to convert serine to EA (4):…”

“…Net synthesis of PtdSerine, which could provide the starting point for the above cycle, can be carried out by the combined actions of phosphatidatecytidylyltransferase [reaction (4) PtdEA is a major phospholipid of higher plants (30,32) and an ethanolamine-phosphotransferase capable of catalzying reaction 9 is well known (25,29,30). However, the cytidylyltransferase catalyzing reaction 8 has been mentioned only very briefly as present in enzyme preparations from carrot root (20), and subsequent attempts in at least two laboratories to demonstrate this activity in extracts of higher plants were unsuccessful (25)(26)(27). Radiolabeled EA is incorporated into PtdEA (1 1,26,27,33,38,39).…”

“…However, the cytidylyltransferase catalyzing reaction 8 has been mentioned only very briefly as present in enzyme preparations from carrot root (20), and subsequent attempts in at least two laboratories to demonstrate this activity in extracts of higher plants were unsuccessful (25)(26)(27). Radiolabeled EA is incorporated into PtdEA (1 1,26,27,33,38,39). In certain systems, the characteristics of this incorporation are such that it was concluded that some or all of it occurred by base exchange (26,27,38), for example, reaction 2 (proceeding from right to left as written above).…”

Complementary DNAs Encoding Eukaryotic-Type Cytidine-5[prime]-Diphosphate-Diacylglycerol Synthases of Two Plant Species

“…Microsomal membranes might account for most of the PI synthesis in plant cells (Moore, 1982). Studies on microsomes isolated from spinach leaves also suggest the presence of phosphatidylserine synthase (EC 2.7.8.8), which utilizes CDP-DG (Marshall and Kates, 1974). Moreover, CDS and other enzymes necessary for PI resynthesis from phospholipase C and D reaction products are present in plant plasma membranes (Wissing et al, 1992).…”