Regulation of CTP:phosphocholine cytidylyltransferase by lipids. 1. Negative surface charge dependence for activation.
The activity of phosphocholine cytidylyltransferase (CT), the regulatory enzyme in phosphatidylcholine synthesis, is dependent on lipids. The enzyme, obtained from rat liver cytosol, was purified in the presence of Triton X-100 [Weinhold et al. (1986) J. Biol. Chem. 261, 5104]. The ability of lipids to activate CT when added as Triton mixed micelles was limited to anionic lipids. The relative effectiveness of the lipids tested suggested a dependence on the negative surface charge density of the micelles. The mole percent lipid in the Triton mixed micelle required for activation decreased as the net charge of the lipid varied from 0 to -2. Evidence for the physical association of CT with micelles and vesicles containing phosphatidylglycerol was obtained by gel filtration. The activation by micelles containing PG was influenced by the ionic strength of the medium, with a higher surface charge density required for activation at higher ionic strength. The micelle surface potential required for full activation of CT was calculated to be -43 mV. A specificity toward the structure of the polar group of the acidic lipids was not apparent. CT was activated by neutral lipids such as diacylglycerol or oleyl alcohol when included in an egg PC membrane, but the activities were reduced by dilution with as little as 10 mol % Triton. Thus Triton mixed micelles are not suitable for studying the activation of CT by these neutral lipid activators. We conclude that one way that lipid composition can control CT-membrane binding and activity is by changing the surface potential of the membrane. Other distinct mechanisms involved in the activation by neutral lipids are discussed.
Cornell
R B
RB
Department of Chemistry/Biochemistry, Simon Fraser University, Burnaby, British Columbia, Canada.
D000818:Animals; D019916:Choline-Phosphate Cytidylyltransferase; D002850:Chromatography, Gel; D003902:Detergents; D004789:Enzyme Activation; D007700:Kinetics; D008081:Liposomes; D008099:Liver / Q000201:enzymology; D008433:Mathematics; D008823:Micelles; D008962:Models, Theoretical; D009713:Nucleotidyltransferases / Q000302:isolation & purification / Q000378:metabolism*; D017830:Octoxynol; D019301:Oleic Acid; D009829:Oleic Acids / Q000494:pharmacology*; D009994:Osmolar Concentration; D010743:Phospholipids / Q000494:pharmacology*; D011092:Polyethylene Glycols; D011485:Protein Binding; D051381:Rats; D013499:Surface Properties
D003902:Detergents; D008081:Liposomes; D008823:Micelles; D009829:Oleic Acids; D010743:Phospholipids; D019301:Oleic Acid; D011092:Polyethylene Glycols; D017830:Octoxynol; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase