CTP:Phosphocholine Cytidylyltransferase Activation by Oxidized Phosphatidylcholines Correlates with a Decrease in Lipid Order:  A ²H NMR Analysis

Abstract

The enzyme CTP:phosphocholine cytidylyltransferase (CT) binds reversibly to membranes and is active only in its membrane-bound form. Membrane lipid composition influences the equilibrium between its soluble and membrane-bound forms. Whereas the enzyme is not activated by phosphatidylcholine (PC) vesicles, it is activated by PC vesicles that have been oxidized with HClO(4) [Drobnies, A. E., et al. (1998) Biochim. Biophys. Acta 1393, 90-98]. Here we explore the mechanism of activation of CT by a PC oxidized with lipoxidase. Multilamellar vesicles (MLVs) containing > or =5 mol % oxidized 1-palmitoyl-2-arachidonoylPC (PAPC) progressively activated the enzyme, which was fully activated by 25 mol % oxidized PC. The effect of oxidized PAPC on lipid order was investigated by (2)H NMR, using MLVs containing PAPC perdeuterated on the palmitoyl chain. Spectral depaking generated order parameter profiles along the sn-1 chain. The average order parameter (S(CD)) in the plateau region at 37 degrees C decreased from 0.18 to 0.15 with increasing percent of oxidized PAPC (0-25%). The change in S(CD) was even greater near the end of the palmitoyl chain. CT activation was inversely related to lipid order. The major component of the lipoxidase-oxidized PAPC was purified and characterized by mass spectrometry and NMR. This component, 1-palmitoyl-2-(11,15-dihydroxy)eicosatrienoylPC (dihydroxyPAPC), incorporated into PAPC MLVs, also stimulated CT activity and reduced the lipid order parameter. Both effects were reversed by egg sphingomyelin. We propose that CT activation by oxidized PAPC is mediated by effects on lipid packing perturbations. This is the first study to report the effects of a purified oxidized PC on the orientational order along the acyl chain and to correlate the lipid disordering of the oxidized PC with the activation of a membrane-associated regulatory enzyme.

Similar gene expression patterns characterize aging and oxidative stress in Drosophila melanogaster

“…Aging and oxidative stress both caused up-regulation of Cct-1, which encodes the rate limiting enzyme in phosphatidylcholine synthesis (33). Cct-1 enzyme is activated by oxidized lipids (34), and Cct-1 gene expression level may affect Drosophila life span (35). Aging and oxidative stress responses also shared the down-regulation of numerous proteasome subunits and other proteases (41 genes), alkaline phosphatases (eight genes), and triacylglycerol lipases (four genes).…”

CTP:phosphocholine cytidylyltransferase: Function, regulation, and structure of an amphitropic enzyme required for membrane biogenesis

“…M domain peptides from rat CCTa [200][201][202] and P. falciparum CCT [203] also bind selectively to anionic lipid Loop L2 75 122 vesicles. Activation of rat CCTa by vesicles containing DAG, unsaturated PEs, oxidized PCs, fatty alcohol, oxysterols, and farnesol has also been reported [184,186,192,193,204,205]. This latter group of lipids can be categorized as type II non-bilayer forming lipids that promote negative curvature strain when mixed with bilayer-stabilizing phospholipids [206].…”

Stearoyl-CoA Desaturase 1 Deficiency Increases CTP:Choline Cytidylyltransferase Translocation into the Membrane and Enhances Phosphatidylcholine Synthesis in Liver

“…3), because mitochondrial GPAT plays a key role in establishing initial asymmetric distribution of saturated and unsaturated FA in PL (1,36) and its deficiency was shown to decrease 16:0 and increase 20:4 contents in hepatic PC and PE (38). Given that insertion of CCT into membrane was shown to be enhanced by polyunsaturated fatty acids (39), increased unsaturated PE level (20,37), and by disordered fatty acyl chains of PL (25,26) in vitro, alterations in PL fatty acid composition and its membrane organization due to Scd1 mutation might directly promote CCT membrane binding. Indeed, recent confocal microscopy studies have revealed that Scd expression is a very potent regulator of membrane domain structure (40).…”

“…One well characterized mode of regulation of CCT activity is through translocation of the protein from an inactive, soluble form to an active, membrane-bound form (20,22). The insertion of CCT into membranes has been shown to increase by the properties of membranes, including loose packing of lipids in the membrane (23), curvature strain (24), and changes in the fatty acid composition of PL (25,26). Because oleic acid is the most abundant monounsaturated fatty acid in mammalian PL, we hypothesized that membrane perturbation that might occur as a consequence of Scd1 deficiency would affect CCT translocation into membranes, and thus SCD might play an important role in regulating PC biosynthesis.…”