Physical properties of the transmembrane signal molecule, sn-1-stearoyl 2-arachidonoylglycerol. Acyl chain segregation and its biochemical implications.
sn-1,2-diacylglycerol (DAG), a key intermediate in lipid metabolism, activates protein kinase C and is a fusogen. Phosphoinositides, the main sources of DAG in cell signaling, contain mostly stearoyl and arachidonoyl in the sn-1 and -2 positions, respectively. The polymorphic behavior of sn-1-stearoyl-2-arachidonoylglycerol (SAG) was studied by differential scanning calorimetry, x-ray powder diffraction, and solid state magic angle spinning (MAS) (13)C NMR. Three alpha phases were found in the dry state. X-ray diffraction indicated that the acyl chains packed in a hexagonal array in the alpha phase, and the two sub-alpha phases packed with pseudo-hexagonal symmetry. In the narrow angle range strong diffractions of approximately 31 and approximately 62 A were present. High power proton-decoupled MAS (13)C NMR of isotropic SAG gave 16 distinct resonances of the 20 arachidonoyl carbons and 5 distinct resonances of the 18 stearoyl carbons. Upon cooling, all resonances of stearoyl weakened and vanished in the sub-alpha(2) phase, whereas arachidonoyl carbons from 8/9 to 20 gave distinct resonances in the frozen phases. Remarkably, the omega-carbon of the two acyl chains had different chemical shifts in alpha, sub-alpha(1), and sub-alpha(2) phases. Large differences in spin lattice relaxation of the stearoyl and arachidonoyl methene and methyl groups were demonstrated by contact time (cross-polarization) MAS (13)C NMR experiments in the solid phases alpha, sub-alpha(1), and sub-alpha(2). This shows that stearoyl and arachidonoyl in SAG have different environments in the solid states (alpha, sub-alpha(1), and sub-alpha(2) phases) and may segregate during cooling. The NMR and long spacing x-ray diffraction results suggest that SAG does not pack in a conventional double layer with the two acyls in a hairpin fashion. Our findings thus provide a physicochemical basis for DAG hexagonal phase domain separation within membrane bilayers.
Hindenes
J O
JO
Department of Biochemistry and Molecular Biology, University of Bergen, Bergen, Norway.
Nerdal
W
W
Guo
W
W
Di
L
L
Small
D M
DM
Holmsen
H
H
D002152:Calorimetry, Differential Scanning; D004075:Diglycerides / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D014961:X-Ray Diffraction
D004075:Diglycerides; C075809:1-stearoyl-2-arachidonoylglycerol