Fig. 3.

LPLA2 activity and LPLA2 protein of the soluble fraction obtained from WT and mutated hLPLA2s gene transfectants. The soluble fractions of individual transfectants were obtained by centrifugation for 1 h at 150,000 g of their cell homogenates (1 mg of protein/ml) at 4°C. A: In LPLA2 assay, 20 μl of each soluble fraction was incubated for 20 min at 37°C with liposomes containing NAS as described in the Material and Methods. B: To obtain the initial velocity of each fraction, the reaction time was adjusted in each assay. Error bars indicate SD (n = 3). C: In Western blotting, 40 μl of each soluble fraction was separated by SDS polyacrylamide gel electrophoresis and subjected to immunoblotting with an anti-hLPLA2 rabbit antibody, and WT and mutated hLPLA2s were visualized as described in the Material and Methods. C and V denote control and expression vector, respectively.