Fig. 7.

Hydrolysis of one medium-chain ox-PC by LPLA2 under neutral conditions. The reaction mixture contained 49 mM HEPES (pH 7.4), 10 μg/ml BSA, liposomes (130 μM PL), and 80 ng/ml of recombinant mouse LPLA2 in 500 μl of total volume. The liposomes consisted of DODPC/DOPC or one medium-chain ox-PC (PAzPC) (2.4:1, molar ratio). The reaction was initiated by the addition of recombinant LPLA2 and kept for 30 to 120 min at 37°C. The reaction products were extracted and separated with a HPTLC plate using a solvent system consisting of chloroform/methanol/pyridine (98:2:0.5, v/v) (A, B) or chloroform/methanol/water (60:35:8, v/v) (C), as described in the Materials and Methods. The reaction products produced by LPLA2 in (A–C) were quantified by scanning the plate, as described in the Materials and Methods, and were plotted against the incubation time (D). 1-Pal-2-LysoPC, 1-palmitoyl-sn-glycero-3-phosphocholine.