FIGURE 4.

Stacked ³¹P NMR spectra from competition assays of three SicTox PLD enzymes with choline- and ethanolamine-containing substrates. Left panels show SM/CPE competition experiments, and right panels show LPC/LPE competition experiments. The spectra illustrate the preference, or lack thereof, of the different enzymes for choline (SM and LPC) or ethanolamine (CPE and LPE) headgroups. Assays contained 10 μg of each SicTox PLD and ∼2 mm of each substrate. In the SM/CPE assay, cleavage of either substrate yields cyclic ceramide phosphate (CCP), a six-membered ring phosphate with an upfield ³¹P resonance. In the LPC/LPE assay, cleavage of either substrate yields cyclic phosphatidic acid (CPA), a five-membered ring phosphate with a far downfield ³¹P resonance.