Functions and biosynthesis of plasmalogens in health and disease

Plasmalogens (1-O-alk-1'-enyl-2-acyl glycerophospholipids) constitute a special class of phospholipids characterized by the presence of a vinyl-ether bond at the sn-1 position. Although long considered as biological peculiarities, interest in this group of phospholipids has grown in recent years, thanks to the realization that plasmalogens are involved in different human diseases. In this review, we summarize the current state of knowledge with respect to the enzymatic synthesis of plasmalogens, the characteristic topology of the enzymes involved and the biological roles that have been assigned to plasmalogens.     

Reductive and oxidative biosynthesis of plasmalogens in myelinating brain

Palmitic acid-1-(14)C and hexadecanol-1-(14)C were administered intracerebrally to 18-day-old rats. Incorporation of radioactivity into the constituent alkyl, alk-1-enyl, and 1-acyl moieties, as well as into the 2-acyl moieties, of the ethanolamine phosphatides of brain was determined after 1, 2, 3, 6, and 22 hr. Incorporation of radioactivity from hexadecanol into both alkyl ethers and alk-1-enyl ethers proceeded at a rate more than 10 times higher than from palmitic acid. Hexadecanol was rapidly oxidized to fatty acids which were incorporated into the acyl moieties of the ethanolamine phosphatides. When palmitic acid was used as a precursor, labeled long-chain alcohols could be isolated from the lipid extract. As labeled long-chain aldehydes could not be detected in any of the lipid extracts, alcohols appear to be key intermediates for the biosynthesis of both alkyl and alk-1-enyl glycerophosphatides.     

Role of dihydroxyacetonephosphate acyltransferase in the biosynthesis of plasmalogens and nonether glycerolipids

The variant CHO-K1 cell line, NRel-4, is unable to synthesize plasmalogens because of a severe reduction in dihydroxyacetonephosphate acyltransferase (DHAPAT) activity (Nagan, N., A. K. Hajra, L. K. Larkins, P. Lazarow, P. E. Purdue, W. B. Rizzo, and R. A. Zoeller. 1998. Isolation of a Chinese hamster fibroblast variant defective in dihydroxyacetonephosphate acyltransferase activity and plasmalogen biosynthesis: use of a novel two-step selection protocol. Biochem. J. 332: 273-279). Northern analysis demonstrated that the loss of this activity was attributable to a severe reduction in mRNA levels for DHAPAT. Transfection of NRel-4 cells with a plasmid bearing the human DHAPAT cDNA recovered DHAPAT activity and plasmalogen biosynthesis. Examination of clonal isolates from the transfected population showed that recovery of as little as 10% of wild-type DHAPAT activity restored plasmalogen levels to 55% of normal, whereas in one isolate, NRel-4.15, which overexpressed DHAPAT activity by 6-fold over wild-type cells, plasmalogen levels were returned only to wild-type values. Although the rate of plasmenylethanolamine biosynthesis was restored in NRel-4.15, the biosynthesis of nonether glycerolipids was either decreased or unaffected, suggesting that peroxisomal DHAPAT does not normally contribute to nonether glycerolipid biosynthesis. These data demonstrate that a defect in the gene that codes for peroxisomal DHAPAT is the primary lesion in the NRel-4 cell line and that the peroxisomal DHAPAT is essential for the biosynthesis of plasmalogens in animal cells.     

The biosynthesis of plasmalogens by rat brain: involvement of the microsomal electron transport system.

The biosynthesis of 1-alk-1'-enyl-2-acyl-sn-glycero-3-phosphoethanolamine (ethanolamine plasmalogens) was studied using 1-[1-14C]hexadecyl-sn-glycero-3-phosphoethanolamine as the substrate and EDTA-washed microsomes from brains of 14-day-old rats. It was found that the 1-E11-14C]hexadecyl-sn-glycero-3-phosphoethanolamine was first acylated to form 1-[1-14C]hexadecyl-2-acyl-sn-glycero-3-phosphoethanolamine, then was desaturated to form 1-[1-14C]hexadec-1'-enyl-2-acyl-sn-glycero-3-phosphoethanolamine. The desaturation required O2 and NADH or NADPH and was inhibited by KCN but not by CO. The data indicated that the desaturation is carried out by a mixed-function oxidase system similar to that involved in the desaturation of fatty acids and that the pathway for the biosynthesis of plasmalogens in brain is similar to that previously found in other tissues. The desaturase was not stimulated by ATP and Mg2plus nor inhibited by EDTA. The specific activity of microsomes from brains of rats of different ages was determined; the activity decreased with age until in adults the activity was only 15% that of the 12--14-day-old rats.     

Metabolism of alkyl glyceryl ethers and their noninvolvement in alkane biosynthesis in plants

[1-¹⁴C]Octadecyl glyceryl ether did not label alkanes in the leaves of Brassica oleracea and Pisum sativum while [1-¹⁴C]octadecanol and [1-¹⁴C]octadecanoic acid readily labeled the alkanes. About 40% of the exogenous-labeled glyceryl ether was incorporated intact into choline phosphatide while 10–20% was converted into fatty acids and alcohols. [1-¹⁴C]octadecanol was not converted into alkyl glyceryl ether, but it was oxidized to the corresponding acid and then incorporated into alkanes. These results show that alkyl ether is not an intermediate in alkane biosynthesis. When [1-¹⁴C-1-³H]-octadecanol was fed to the leaves of B. oleracea and P. sativum, only the ¹⁴C and no ³H was incorporated into alkanes, ketones, and secondary alcohols. These results show that fatty alcohols are first oxidized to the acid before being incorporated into alkanes, ruling out fatty alcohol, alkyl ether, and alk-1-enyl ether as intermediates in alkane biosynthesis. The exogenous alcohols were also readily esterified into wax esters in both tissues.     

Deuterium nuclear magnetic resonance studies on the plasmalogens and the glycerol acetals of plasmalogens of Clostridium butyricum and Clostridium beijerinckii

Deuterium nuclear magnetic resonance was used to investigate the structure of different lipid fractions isolated from the anaerobic bacteria Clostridium butyricum and Clostridium beijerinckii. The fractions isolated from C. butyricum were (1) phosphatidylethanolamine/plasmenylethanolamine and (2) the glycerol acetal of plasmenylethanolamine, and from C. beijerinckii similar fractions containing principally (1) phosphatidyl-N-monomethylethanolamine, along with its plasmalogen, and (2) the glycerol acetal of this plasmalogen were isolated. The third fraction from both species consisted largely of the acidic lipids phosphatidylglycerol and cardiolipin along with plasmalogen forms of these lipids. Palmitic acid with deuterium labels at C-2, C-3, or C-4 or oleic acid with deuterium labels at C-2 and C-9,10 was added to the growth medium and incorporated to various extents in the lipid fractions. Biochemical analysis showed that palmitic acid and oleic acid were preferentially bound to the sn-2 and sn-1 positions, respectively, of the glycerol backbone when both fatty acids were added to the medium. From the 2H NMR spectra, the hydrocarbon chain ordering near the lipid-water interface could be determined and appeared to be similar for all three lipid fractions. The deuterium quadrupole splitting and order parameter were low at the C-2 segment and increased by almost a factor of 2 at positions C-3 and C-4 for cells fed with deuterated palmitic acid along with unlabeled oleic acid. These results agree with previous findings on pure diacyl lipids in which the sn-2 chain was found to adopt a bent conformation at the carbon segment C-2. However, two unusual quadrupole splittings could be detected for the plasmalogens.(ABSTRACT TRUNCATED AT 250 WORDS)     

The cerebro-hepato-renal (Zellweger) syndrome. Impaired de novo biosynthesis of plasmalogens in cultured skin fibroblasts

In tissues of patients with the cerebro-hepato-renal (Zellweger) syndrome the plasmalogen content is very low. In order to study the biosynthesis of plasmalogens, skin fibroblasts of Zellweger patients, controls and heterozygotes, and amniotic fluid cells of controls were cultured in a medium supplemented with [1-14 C]hexadecanol or 1-O-[9,10-3H2]octadecylglycerol. The incorporation of 14C-label into the alkenyl moiety of plasmalogens was strongly reduced in Zellweger patients as compared to controls. The low concentration of 14C-labeled plasmalogens was not compensated for by an elevated levels of 14C-labeled alkyl phospholipids. Hexadecanol was partly oxidized to fatty acid in all cell lines and the incorporation of 14C-labeled fatty acid into phospholipids was comparable for patients and controls. [3H]Alkylglycerol was incorporated into plasmalogens with the same efficiency in Zellweger patients as in controls. These results indicate that only the reaction(s) involved in the introduction of the ether bond in the process of plasmalogen synthesis are deficient in Zellweger patients. The results also suggest that the hexadecanol incorporation patterns can be used for the (prenatal) diagnosis of the Zellweger syndrome.     

Lipidomic analysis of bacterial plasmalogens

Plasmalogens are a group of lipids with potentially important, and not yet fully known, functions in organisms from bacteria to protozoans, invertebrates, and mammals. They can protect cells against the damaging effects of reactive oxygen species, protect other phospholipids or lipoprotein particles against oxidative stress, and have been implicated as signaling molecules and modulators of membrane dynamics. They have been found in many anaerobic bacterial species, and their biosynthetic pathways differ in aerobic and anaerobic organisms. The use of advanced techniques permits the identification of not only plasmalogen classes but also their positional isomers and often also individual molecular species. This paper describes direct analyses of plasmalogens from natural sources, frequently very unusual, using electrospray ionization mass spectrometry in combination with high-performance liquid chromatography and/or shotgun lipidomics.