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. Author manuscript; available in PMC: 2021 Nov 1.
Published in final edited form as: Biochim Biophys Acta Mol Cell Biol Lipids. 2021 Jul 29;1866(11):159017. doi: 10.1016/j.bbalip.2021.159017

Fig. 3.

Fig. 3.

CPE synthase, SMS, PE-PLC, PC-PLC, PS-PLC, PG-PLC, and PAP activity assays, using recombinant SMSr-flag (rSMSr) and exogenous substrates. Purified rSMSr (150 ng) was used for enzyme assays. (A) CPE synthase activity assay, using NBD-Cer and PE as two substrates. (B) SMS activity assay, using NBD-Cer and PC as two substrates. (C) PE-PLC activity assay, using NBD-PE as a substrate. Different amount of rSMSr were used. (D) PE-PLC activity assay, using 14C-PE as a substrate. (E) PC-PLC activity assay, using 14C-PC as a substrate. (F) PS-PLC activity assay, using NBD-PS as a substrate. (G) PG-PLC activity assay, using NBD-GC as a substrate. (H) PAP, PE-PLC, and PC-PLC activity assays, using NBD-PA, NBD-PE, and NBD-PC as a substrate, respectively. −, no protein was added. +, Bacterial PLC was used as a positive control. The development solvent for (A) and (B) was Chloroform:Methanol:NH4OH: 14:6:1 (v/v/v); The development solvent for (C)–(G) was Chloroform: Methanol: 15:1 (v/v). The results were the representative of three independent experiments.