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Cytidine Diphosphate Diacylglycerol

Nucleotides are the basic building blocks of DNA and RNA, and they are required for many aspects of intermediary metabolism. They consist of three elements – a heterocyclic nitrogenous base derived from a purine (adenine and guanine) or pyrimidine (uracil, thymine and cytosine), a pentose (ribose or deoxyribose) and phosphoric acid. Cytidine, for example, consists of cytosine attached to a ribofuranose ring via a β-N1-glycosidic bond. From the standpoint of lipid metabolism, one of the most important of the nucleotide metabolites is cytidine 5’-phosphoric acid, which is a key component of the phospholipid cytidine diphosphate diacylglycerol (CDP-diacylglycerol or CDP-DAG), a liponucleotide. This lipid was first found in eukaryotic organisms, but it is now known to be ubiquitous and has an essential role in the biochemistry of prokaryotes also as an intermediate in the biosynthesis of many complex glycerolipids. Its discovery owes much to serendipity allied to the receptive mind of the American biochemist Eugene Kennedy (see our web page on phosphatidylcholine).

Structural formula of cytidine diphosphate diacylglycerol

Cytidine diphosphate diacylglycerol per se is hardly ever noticed in analyses of lipid compositions of tissues, as it is present is such small amounts, perhaps only 0.05% or so of the total phospholipids, and as a consequence, the composition of the fatty acids or molecular species in this lipid in nature (as opposed to experiments in vitro) is rarely reported. Data for ox brain are listed in Table 1. In this instance, the composition is closer to that of phosphatidylinositol for which it is a precursor than to that of any other lipid.

Table 1. Fatty acid compositions of positions sn-1 and sn-2 of the cytidine diphosphate diacylglycerol of bovine brain.
Fatty acids
16:0 18:0 18:1 18:2 20:4 22:3 22:6
sn-1 15 78 6
sn-2 4 10 35 3 45 3 1
Thompson, W. and MacDonald, G. Eur. J. Biochem., 65, 107-111 (1976).

Biosynthesis: Biosynthesis of CDP-diacylglycerol (CDP-DAG) involves condensation of phosphatidic acid (PA) and cytidine triphosphate, with elimination of pyrophosphate, catalysed by an enzyme CDP-diacylglycerol synthase (CDS). This is an membrane protein and contains six transmembrane domains in eukaryotes. Two different isoforms of the enzyme associated with the outer surface of the endoplasmic reticulum and with differing tissue distributions have been found in animals, of which one (CDS2) is expressed ubiquitously and is selective for the acyl chains at the sn-1 and sn-2 positions and for 1-stearoyl-2-arachidonoyl-sn-phosphatidic acid in particular; this may be a distinct pool utilized for phosphatidylinositide synthesis. The second isoform (CDS1) has a more restricted pattern of expression and is most active in the heart. It has no special substrate specificity and may create a separate pool of CDP-diacylglycerols for other purposes. Mutant mammalian cells lacking either CDS1 or CDS2 expression produce giant or super-sized lipid droplets in culture but by different mechanisms, though excessive phosphatidic acid production may be a factor by causing fusion of smaller lipid droplets.

Biosynthesis of cytidine diphosphate diacylglycerol

A single iso-enzyme is present in yeasts and bacteria, but five forms are found in plants such as Arabidopsis, i.e. CDS1, CDS2, and CDS3 in the endoplasmic reticulum and CDS4 and CDS5 in the chloroplasts. In yeast and animals, the loss of CDS results in lethality, but in plants a single knock-out of CDS2, CDS3, CDS4 or CDS5 did not exhibit any visible phenotype. However, double mutants of the plant genes caused death at an early stage in seedling growth.

The CDS from the hyperthermophilic bacterium Thermotoga maritima crystallized as a dimer, with each subunit having a deep funnel-shaped cavity containing a catalytically important Mg2+,K+-hetero-di-metal center and open both to the cytoplasmic side of the bilayer and to the membrane. Presumably, the water-soluble and lipidic substrates (CTP, PA) and products (pyrophosphate, CDP-DAG) can enter and exit this cavity, and phosphatidic acid could potentially enter this from either the inner (cytoplasmic) or outer leaflet of the membrane. The CDP-DAG product of the reaction probably exits on the cytoplasmic side.

A further distinct CDP-diacylglycerol synthase (translocator assembly and maintenance protein 41 or TAM41), first characterized in yeast, is located in the inner membrane of mitochondria facing the matrix, while a peripheral mitochondrial protein (TAMM41) is now known to be the mammalian equivalent. This uses phosphatidic acid synthesised in the endoplasmic reticulum and transported into mitochondria, presumably with the aid of the contact region between the endoplasmic reticulum and mitochondria known as the mitochondria-associated membrane; the CDP-diacylglycerol formed by these enzymes is used mainly for cardiolipin synthesis. In the protozoan parasite, Toxoplasma gondii, there are two CDP-diacylglycerol synthases of which one is a eukaryotic type in the endoplasmic reticulum (TgCDS1) and the second a prokaryotic type (TgCDS2) in the apicoplast, a non-photosynthetic plastid-like organelle. Two discrete CDP-DAG pools are produced for the subsequent synthesis of phosphatidylinositol from the first in the Golgi bodies and phosphatidylglycerol from the second in the mitochondria.

Function: CDP-diacylglycerol produced by these enzymes is utilized immediately for the synthesis of phosphatidylglycerol (PG) and phosphatidylinositol (PI) in the endoplasmic reticulum, and of cardiolipin via the intermediate phosphatidylglycerol in mitochondria. Turnover is very rapid and the pool of CDP-diacylglycerol is always much smaller than that of the precursor phosphatidic acid. In animals, phosphatidylcholine (PC), phosphatidylethanolamine (PE) and triacylglycerols (TAG) are synthesised via the Kennedy pathway mainly with diacylglycerols as the key intermediate, and this is also true for monogalactosyldiacylglycerols (MGDG) in plants.

CDP-DG and Kennedy pathways of lipid synthesisIn fungi and prokaryotes, CDP-diacylglycerol is also the precursor for phosphatidylserine. In yeast such as Saccharomyces cerevisiae, CDP-diacylglycerol is one of the precursors for phosphatidylethanolamine, which can in turn be converted via mono- and dimethylphosphatidylethanolamines (PME and PDME) to phosphatidylcholine, although the Kennedy pathway also functions. In the bacterium, Escherichia coli, CDP-diacylglycerols with both ribose and deoxyribose as the sugar component are produced, and both are utilized as substrates by phosphatidylserine and phosphatidylglycerophosphate synthases (no synthesis occurs via the Kennedy pathway with diacylglycerol intermediates).

It is not known whether the final fatty acid composition of the lipid is a result of the specificity of the CDP-diacylglycerol synthase in selecting particular molecular species of phosphatidic acid, or whether remodelling occurs via deacylation/re-acylation reactions as in the Lands' cycle prior to synthesis of other lipids, apart from the action of CDS2 in the biosynthesis of phosphoinositides.

Most studies of CDP-diacylglycerol have been concerned with its function as an intermediate in the biosynthesis of other lipids, and as such this is the first step in a pathway that leads only indirectly to phosphatidylethanolamine and phosphatidylcholine. In the main biosynthetic pathways to these lipids, nucleotides (CDP-ethanolamine and CDP-choline) are required but not liponucleotide intermediates. Similarly, another nucleotide, i.e. uridine 5-diphosphate(UDP)-hexose (where hexose = glucose, galactose, etc), is required for the formation of glycolipids, including both the glycosyldiacylglycerols and sphingoglycolipids.

The extent of the biological functions of CDP-diacylglycerol, other than as an intermediate in phospholipid biosynthesis, is only partly understood. However, CDP-diacylglycerol synthase is also a regulator of phospholipid metabolism, as it is believed to be the rate-limiting enzyme in phosphatidylinositol biosynthesis. In consequence, it has a role in the regulation of lipid-dependent signal transduction processes. It also has a function in controlling the size of lipid droplets in adipocytes, presumably by regulating the concentration of phosphatidic acid. Anti-cancer activities in vitro have been reported.

Analysis: Because it is such a minor component of tissues, isolation of cytidine diphosphate diacylglycerol appears to be a tedious task, and the only detailed published analysis of which I am aware involved ion-exchange column chromatography and thin-layer chromatography. The pyrophosphate bond is relatively labile and is very susceptible to hydrolysis, especially under basic conditions. At natural tissue concentrations of this lipid, even the more modern mass spectrometric methods do not appear to be sufficiently sensitive.

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Lipid listings Credits/disclaimer Updated: November 8th, 2019 Author: William W. Christie LipidWeb icon