Biochemistry of polyglycerophosphatides in central nervous tissue. I. On the biosynthesis, structure, and enzymatic degradation of phosphatidylglycerophosphate and phosphatidylglycerol in isolated sheep brain mitochondria

Abstract

The formation of labelled phosphatidylglycerophosphate and phosphatidylglycerol from L-glycero-3-phosphate-2-3H and cytidine diphosphate (CDP)-D-diglyceride in subcellular particles of sheep brain has been studied, establishing the predominant location of this enzyme system in mitochondria. The general characteristics and the optimal conditions for the biosynthesis of these lipids have been determined and the chromatographic separation and isolation of phosphatidylglycerophosphate and phosphatidylglycerol are described. In addition, the chemical and enzymatic degradations supporting the structure and configuration for each biosynthesized lipid are presented. The enzymatic dephosphorylation of phosphatiadylglycerophosphate to phosphatidylglycerol with mitochondria isolated from sheep brain has been found. No biosynthesized cardiolipin was, however, detected.The mechanism of the enzymatic reactions for the formation of these lipids has been studied by using L-glycero-3-phosphate[33P]-2-3H and CDP-D-diglyceride; the pathway found by Kennedy et al. (1) for the biosynthesis of phosphatidylglycerol in liver is applicable in the biosynthesis of this compound in sheep brain mitochondria.

Inter-Relations between the Phospholipids of Rat Pancreatic Islets during Glucose Stimulation and Their Response to Medium Inositol and Tetracaine

“…In addition, it is apparent that the build-up of CDP diglyceride diverts more labelling into phosphatidylglycerol, presumably by reaction with glycerol phosphate and subsequent dephosphorylation (Scheme 1). The enzymes catalysing the last reactions are thought to be confined to mitochondria at least in mammalian tissues [21,22]. Although phosphatidylglycerol is an intermediary in cardiolipin biosynthesis the labelling of this latter phospholipid was minimal with the incubation times used in the present experiments.…”

Enzymatic degradation and partial biosynthetic reconstitution of microsomal and mitochondrial membranes

“…Methods describing thin-layer and paper chromatography were previously described (7). Mild alkaline hydrolysis of lipids and chromatography of the water-soluble products was previously described ('7)-Determination of radioactivity using a Packard Tri-Carb scintillation spectrometer 3W3 was described (7).…”

“…Mild alkaline hydrolysis of lipids and chromatography of the water-soluble products was previously described ('7)-Determination of radioactivity using a Packard Tri-Carb scintillation spectrometer 3W3 was described (7). Phosphoms was determined according to Bartlett (81, and esters according to Stein and Shapiro (91 Hncubutisns sf Membranes with Radioactive Substrates g=H)P-fmethyHCC]Choline-The reaction mixture contained 5.0 mg of microsoma1 or EO rng of mitochondria1 membrane protein, 2.0 r*mol s f CDP-Emetday8-14C]choline (SA = 7 x lo5 eprn/pmol), 5 pmol of L-eysteine, 3 pmol of ATP, and 75 pmol of Tris-HCl bufier (pH 7.4) in a final volume of 0-5 rnE.…”

Biosynthesis of microsomal phospholipids and mitochondrial polyglycerophosphatides in rapidly sedimenting endoplasmic reticulum

“…Furthemore, in our experiments, both cmde md purified preparations of RSER were examined and compared with microsomes and cmde md purified mitochon~a. Results of these experiments are shown in Table 1 , and an diquot was analyzed for radioactivity and composition by paper chromatography of water-soluble compounds obtained after mild alkaline hydrolysis of Iipids as described (12). Mitochondria and RSER were purified by ultracentrifugation through a sucrose gradient as described in Materids and methods.…”