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Arch Biochem Biophys. 1998 May 1;353(1):131-40.

Substrate selectivities and lipid modulation of plant phospholipase D alpha, -beta, and -gamma.

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Department of Biochemistry, Kansas State University, Manhattan, Kansas 66506, USA.


Three classes of phospholipase D (PLD), designated PLD alpha, -beta, and -gamma, have been cloned from plants, but their substrate selectivities have not been established. Using active PLDs expressed from their cDNAs in Escherichia coli, we compared the hydrolytic activities of these three PLDs toward various phospholipids and the influence of substrate composition on their substrate selectivities. When single-class phospholipid vesicles of phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylserine (PS), phosphatidylinositol (PI), phosphatidylinositol 4,5-bisphosphate (PIP2), N-acylphosphatidylethanolamine (NAPE), and cardiolipin (CL) were examined, PLD alpha hydrolyzed PC, PE, and PG but PLD beta and -gamma showed no activity toward any of these lipids. When PIP2 was included in mixed vesicles with the phospholipids above, PLD alpha showed the same PC-, PE-, and PG-hydrolyzing ability, whereas PLD beta and -gamma were able to hydrolyze both PE and PS. When both PE and PIP2 were included in substrate vesicles, PLD beta and PLD gamma hydrolyzed PC, PG, and NAPE, showing that both PE and PIP2 are required for PC, PG, and NAPE hydrolysis by PLD beta and -gamma. The PE activation of PLD beta and -gamma required lipid vesicles made of mostly PE, suggesting that PE may affect the substrate presentation rather than serve as a cofactor of these PLDs. Under equivalent reaction conditions, PLD beta displayed a similar preference for PC and NAPE, whereas PLD gamma preferred NAPE to PC by nearly three times. None of the three PLDs used PI, CL, or PIP2 as substrates. These results have identified PS- and NAPE-hydrolyzing PLDs and have indicated an important role for lipid composition in regulating the substrate selectivity of PLD beta and -gamma.

[Indexed for MEDLINE]

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