Physical Properties of the Transmembrane Signal Molecule, sn-1-Stearoyl 2-Arachidonoylglycerol

• Published in: The Journal of biological chemistry 2000-03, Vol.275 (10), p.6857-6867
• Main Authors: Hindenes, Jan-Ove, Nerdal, Willy, Guo, Wen, Di, Li, Small, Donald M., Holmsen, Holm
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.275.10.6857

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 thesn-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) 13C NMR. Three α phases were found in the dry state. X-ray diffraction indicated that the acyl chains packed in a hexagonal array in the α phase, and the two sub-α phases packed with pseudo-hexagonal symmetry. In the narrow angle range strong diffractions of ∼31 and ∼62 Å were present. High power proton-decoupled MAS 13C 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-α2 phase, whereas arachidonoyl carbons from 8/9 to 20 gave distinct resonances in the frozen phases. Remarkably, the ω-carbon of the two acyl chains had different chemical shifts in α, sub-α1, and sub-α2 phases. Large differences in spin lattice relaxation of the stearoyl and arachidonoyl methene and methyl groups were demonstrated by contact time (cross-polarization) MAS 13C NMR experiments in the solid phases α, sub-α1, and sub-α2. This shows that stearoyl and arachidonoyl in SAG have different environments in the solid states (α, sub-α1, and sub-α2phases) 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.