Conversion of Phosphatidylcholine to Phosphatidylglycerol with Phospholipase D and Glycerol
2007
Piazza, George J. | Marmer, William N.
When phosphatidylcholine (PtdCho) is acted upon by the enzyme phospholipase D (PLD) in the presence of glycerol and water, at least two products can arise: phosphatidic acid (PtdOH) from hydrolysis and PtdGly from transphosphatidylation. Commercial PLD preparations from Streptomyces chromofuscus, Streptomyces sp., and cabbage were examined for their ability to selectively promote PtdGly formation in a two phase aqueous-organic solvent system. Factors examined were enzyme amount, pH, glycerol concentration, and the type of organic solvent. The reaction of PtdCho to give products such as PtdGly was followed by HPLC using a stationary phase consisting of a polymerized poly (vinyl alcohol) on silica gel with ELSD. The identities of all products were confirmed by retention times and HPLC-MS analyses. Under all tested conditions PLD from S. chromofuscus gave at most a 15% yield of PtdGly. Higher amounts of added glycerol inhibited this PLD. Nearly quantitative conversion to PtdGly was obtained with cabbage PLD when the mol ratio of glycerol to PtdCho was at least 64 (mol water/glycerol = 105). With PLD from Streptomyces sp. a nearly quantitative yield of PtdGly was obtained when the mol ratio of glycerol to PtdCho was at least 5.3 (mol water/glycerol = 1,266), demonstrating that this PLD had a higher selectivity for glycerol than cabbage PLD. When the glycerol concentration was very low, the level of PtdOH increased, and cardiolipin (diphosphatidylglycerol) was generated. The highest mol ratio PtdGly to PtdOH was observed when the solvents isopropyl ether or dichloromethane were used.
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