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Phosphonolipids



Phosphonolipids consist of 2-aminoethylphosphonic acid (ciliatine) residues attached to a lipid backbone, which can be either a either a ceramide, diacylglycerol or even a carbohydrate moiety of a glycolipid, i.e. the lipids have a carbon-phosphorus bond rather than carbon-oxygen-phosphorus bonds. Lipid-bound aminoethylphosphonic acid was first detected in the single-celled microorganism Tetrahymena pyriformis and then in protozoa. This proved to be a glycerophosphonolipid, but ceramide 2-aminoethylphosphonate, first found in sea anemones, and related sphingolipids are more often encountered and have been more studied.


1. Glycerophosphonolipids

The first of the phosphonolipids to be definitively characterized was a phosphono analogue of phosphatidylethanolamine, i.e. the glycerophosphonolipid 1,2-diacyl-sn-glycerol-3-(2'-aminoethyl)phosphonate ('phosphonylethanolamine'), which is the main phosphonolipid in T. pyriformis. It has also been found in several more species of protozoa, and at low levels in some plant species, various bovine tissues and even in human aorta, and it can exist in diacyl, alkylacyl and alkenylacyl forms. In addition, related N-methyl and N,N-dimethyl phosphonolipids have been detected, together with phosphono analogues of phosphatidylcholine, phosphatidylglycerol and phosphatidylserine.

Formula of 1,2-diacyl-sn-glycerol-3-(2'-aminoethyl)phosphonate

In the phosphonylethanolamine of T. pyriformis, which exists only in the diacyl form, C16 and C18 fatty acids with one to three double bonds predominate with the unsaturated fatty acids concentrated in position sn-2, as listed in Table 1.

Table 1. Positional distributions of fatty acids (mol %) in the 1,2-diacyl-sn-glycerol-3-(2'‑aminoethyl)-phosphonate of T. pyriformis grown at 39.5°C
Fatty acid
14:0 16:0 9-16:1 17:0 9-18:1 6,11-18:2 9,12-18:2 18:3(n-6)
 
sn-1 26 39 8 3 1 1 1 4
sn-2 2 3 12 5 7 8 12 45
Data from Watanabe, T., Fukushima, H. and Nozawa, Y. Biochim. Biophys. Acta, 620, 133-140 (1980).

In plants (Kenaf and cotton seeds), the main fatty acids in the phosphonylethanolamine are saturated and monoenoic, largely 16:0 and 18:1.


2. Ceramide 2-Aminoethylphosphonate and Related Sphingolipids

Ceramide 2-aminoethylphosphonate has been characterized in a wide variety of marine organisms including many invertebrates, and especially ciliated protozoa, coelenerates, gastropods, corals and bivalves, sometimes accompanied by small amounts of N-methylaminoethyl, N,N-dimethylaminoethyl and choline analogues, and it appears to be the most widespread phosphonolipid in nature. In jellyfish, ceramide 2-aminoethylphosphonate is concentrated in the membranes of the tentacles (oral arms) containing the stinging cells, where it may offer resistance to hydrolysis by the endogenous phospholipase A2. Phosphonolipids have also been observed in plants, bacteria and several vertebrates, including humans, although with the last they almost certainly originate from dietary sources and are not synthesised de novo.

Formula of ceramide 2-aminoethylphosphonate + hydroxy form

In marine invertebrates, in addition to sphingosine, sphingadiene and other dihydroxy bases, the ceramide component can contain appreciable amounts of trihydroxy bases depending on the species and tissue; a unique trienoic sphingoid base, 2-amino-4,8,10-octatriene-1,3-diol (d18:3) and a 9-methylated form are sometimes detected. In the phosphonolipids of the marine invertebrate Anthopleura elegantissima, palmitic acid comprises 80% of the total, while 2-hydroxy fatty acids only are found in the corresponding lipids of Pinctada martensii; again the relative proportions of the two fatty acid forms depend on species and tissue. The soft coral Xenia sp. consists largely of a single molecular species with palmitic acid linked to a dienoic sphingoid base.

Ceramide 2-aminoethylphosphonate is present in the membranes of the Gram-negative myxobacterium Sorangium cellulosum (together with the phosphoryl analogue); unexpectedly, the sphingoid base component is one typical of fungi. In addition, phosphonolipids with 1-hydroxy-2-aminoethane attached to the phosphorus moiety have been found in some bacterial species. For example, Bacteriovorax stolpii strain UKi2, a facultative predator-parasite of larger Gram-negative bacteria, synthesises sphingophosphonolipids with this novel head group. The long-chain base components in this instance are mainly C17 iso-methyl-branched phytosphingosine and iso-branched dihydrosphingosine, while the N-linked fatty acids are iso-methyl branched usually with a 2-hydroxyl group. This organism also contains sphingolipids with a 2-amino-3-phosphonopropanate head group.

Further phosphonoglycosphingolipids, i.e. with 2-aminoethylphosphonic units attached to carbohydrate moieties, such as 6-O-(aminoethylphosphono)-galactosyl ceramide and its N-methylethane analogue, related oligoglycosphingolipids, and even a triphosphonoglycosphingolipid have been found in marine invertebrates such as the mollusc, Aplysia kurodai. This organism lacks gangliosides, but complex oligoglycosphingolipids with both phospho- and phosphonoethanolamine groups attached appear to take their place. Similar glucose-containing phosphonolipids and others with the aminoethylphosphonate group on position 4 of the glucose unit have been characterized from a crab.

Formula of 6-O-(aminoethylphosphono)galactosyl ceramide


3. Other Phosphonolipids

An unusual biosurfactant, 2-acyloxyethylphosphonate, has been isolated from waterblooms of the cyanobacterium Aphanizomenon flos-aquae; palmitic acid comprised 80% of the fatty acid components of the lipid, and it was accompanied by some trienoic acids.

Formula of 2-acyloxyethylphosphonate

Phosphonoethanolamine residues have been reported as components of the lipophosphonoglycans of the protozoan parasite Acanthamoeba castellanii, but the point of attachment is not known.


4. Metabolism of Phosphonolipids

More than one pathway for the biosynthesis of 2-aminoethylphosphonate is known, but the simplest requires three enzymes and utilizes phosphoenolpyruvate as the key precursor as illustrated. Little appears to be known of how it is then incorporated into lipids, although in rat hepatocytes, it is incorporated into phosphonolipid by a similar pathway to that for phosphatidylethanolamine biosynthesis.

Biosynthesis of 2-aminoethylphosphonate - the phosphonolipid precursor

While there has been some speculation, little is known of the function of phosphonolipids. They are presumed to be membrane constituents, and they are known to be resistant towards the action of endogenous phospholipases. For example, in Tetrahymena, phosphatidylethanolamine turns over much more rapidly than the phosphono analogue. Therefore, they may have a role in protecting cell membranes from attack by enzymes or from harsh environmental conditions. On the other hand, a number of routes to cleavage of the carbon-phosphorus bond have been identified, including the action of specific lyases and an oxidative mechanism.

A study of the fate of dietary ceramide 2-aminoethylphosphonate in mice demonstrated that sphingoid bases and ceramides appeared rapidly in the mucosa of the small intestine possibly via the action of alkaline sphingomyelinase; it is not yet known what happens to the carbon-phosphorus bond.


5. Analysis

Analysis of phosphonolipids presents no particular problems, except when they co-exist with the conventional phosphate forms of the lipids, which have very similar chromatographic properties. However, methods of isolation have been devised even in these difficult circumstances (see the references cited below). The carbon-phosphorus bond is not hydrolysed by such harsh chemical treatments as boiling in strong acid or base. 31P-Nuclear magnetic resonance spectroscopy is invaluable for detecting the presence of phosphonolipids in lipid extracts, while electrospray-ionization tandem mass spectrometry now appears to hold particular promise for structural analyses.


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Lipid listings Credits/disclaimer Updated: May 23rd, 2017 Author: William W. Christie LipidWeb icon