1-2-oleoylphosphatidylcholine and cholesteryl-sulfate

Topics: Cholesterol Esters; Enzyme Activation; Humans; Kinetics; Liposomes; Phosphatidylcholines; Phospholipases A; Phospholipases A2; Protein Binding; Spectrometry, Fluorescence

The extracellular concentration of the Group II human non-pancreatic secreted phospholipase A2 (hnpsPLA2) is elevated in a variety of inflammatory disorders. This enzyme is remarkable because it demonstrates almost zero activity with egg phosphatidylcholine (PC) or synthetic dioleoyl-phosphatidylcholine (DOPC) as substrate, but expresses high activity with the anionic phospholipid dioleoyl-phosphatidylglycerol (DOPG), a feature shared with the Group II enzyme from rat liver. The presence of certain membrane-bound anions can enhance hydrolysis of PC by the mammalian secreted PLA2S. In this study the ability of various non-polar anions to stimulate DOPC hydrolysis by secreted PLA2S has been investigated. The naturally occurring membrane anion, cholesterol sulphate, was particularly effective in stimulating the hydrolysis of both DOPC and also 1-stearoyl-2-arachidonyl phosphatidylcholine by hnpsPLA2. Activation of DOPC hydrolysis was also achieved with dioleoyl-phosphatidylserine (DOPS); however, DOPS was less effective than cholesterol sulphate. In contrast, the dianion dioleoyl-phosphatidic acid, a known activator of pig pancreatic PLA2, failed to activate the human enzyme. It remains to be established whether cell plasma-membrane hydrolysis by extracellular hnpsPLA2 can be activated in vivo by the presence of suitable membrane anions such as cholesterol sulphate and thus promote an inflammatory response within the cell.

Topics: Amino Acid Sequence; Animals; Anions; Bile Acids and Salts; Cell Membrane; Cholesterol Esters; Humans; Liver; Molecular Sequence Data; Pancreas; Phosphatidic Acids; Phosphatidylcholines; Phosphatidylglycerols; Phosphatidylserines; Phospholipases A; Phospholipases A2; Rats; Swine

One- and two-dimensional 31P-exchange NMR has been used to investigate chemical exchange between coexisting lamellar (L alpha) and non-lamellar (hexagonal HII and cubic I2) lipid phases. Samples of DOPE, DOPE/DOPC (9:1 and 7:3), DOPE/cholesterol sulfate (9:1), DOPC/monoolein (MO) (3:7 and 1:1), and DOPC/DOPE/cholesterol (1:1:2) were macroscopically oriented on glass plates and studied at the 0 degree orientation (angle between the bilayer normal and the external magnetic field), where the L alpha, HII, and I2 resonances are resolved. A reversible L alpha to HII transition was observed for all of the samples except for the DOPC/MO mixtures, which displayed a reversible L alpha to I2 transition. Near-equilibrium mixtures of L alpha and either HII or I2 were obtained after prolonged incubation at a given temperature. Two-dimensional exchange experiments were performed on DOPE at 9-14 degrees C for mixing times ranging from 500 ms to 2 s. For all samples, one-dimensional exchange experiments were performed for mixing times ranging from 100 ms to 4 s, at temperatures ranging from 3 degrees C to 73 degrees C. No evidence of lipid exchange between lamellar and non-lamellar phases was observed, indicating that if such a process occurs it is either very slow on the seconds' timescale, or involves an undetectable quantity of lipid. The results place constraints on the stability or kinetic behaviour of proposed transition intermediates (Siegel, D.P. (1986) Biophys. J. 49, 1155-1170).

Topics: Chemical Phenomena; Chemistry, Physical; Cholesterol Esters; Glycerides; Magnetic Resonance Spectroscopy; Phosphatidylcholines; Phosphatidylethanolamines; Temperature