Effect of temperature on the stereoselectivity of phospholipase D toward glycerol in the transphosphatidylation of phosphatidylcholine to phosphatidylglycerol

Abstract

In this study, the effect of temperature on the stereoselectivity of phospholipase D (PLD) toward the two primary hydroxyl groups of glycerol in the transphosphatidylation reaction of phosphatidylcholine to phosphatidylglycerol (PtdGro) was investigated. For this purpose, PLD from bacteria (Streptomyces septatus TH-2, S. halstedii subsp. scabies K6, and Actinomadura sp.) and cabbage were tested. At the reaction temperatures employed (0-60 degrees C), the proportions of the two PtdGro diastereomers, namely, 1,2-dioleoyl-sn-glycero-3-phospho-3'-sn-glycerol (R,R configuration) and 1 ,2-dioleoyl-sn-glycero-3-phospho-1'-sn-glycerol (R,S configuration), which were produced with PLD from Streptomyces TH-2 and Actinomadura sp., changed gradually from 50% R,R and 50% R,S at 50-60 degrees C to 70% R,R and 30% R,S at 0 degrees C. These alterations suggested that the stereoselectivity of the bacterial PLD toward the two primary hydroxyl groups of prochiral glycerol was significantly influenced by reaction temperature. PLD from Streptomyces K6 showed relatively little effect of temperature on stereoselectivity, giving 65-69% R,R in the temperature range of 60-10 degrees C examined. The plots of In ([R,R]/[R,S]) vs. 1/T gave good linear fits for these three bacterial PLD. No temperature effect was observed for cabbage PLD, which gave an almost equimolar mixture of the R,R and R,S diastereomers in the range from 0 to 40 degrees C. The temperature-dependent change in enantiomeric selectivity of the bacterial PLD promises potentially profitable commercial exploitation.

Simple synthesis of diastereomerically pure phosphatidylglycerols by phospholipase D‐catalyzed transphosphatidylation

“…This study emphasizes the diastereomeric purity of the products produced by PLD-catalyzed transphosphatidylation. Only recently has it become possible to establish the diastereomeric nature of the transphosphatidylation products of various PLD enzymes and to characterize their relaxed substrate specificity (20,26). In an earlier study (16), the diastereomeric nature of the products was implied from the starting materials (isopropylideneglycerols), which were not enantiomerically characterized or their source indicated.…”

Development and application of chromatographic methods for glycerolipid analysis

“…For this purpose, 1,2-O -isopropylidene PtdGro diastereomers were prepared from 1,2-diacyl-sn-glycero-3-phosphocholine (PtdCho) and enantiomeric 1,2-O -and 2,3-O -isopropylidene-snglycerols [51] by transphosphatidylation with phospholipase D from Actinomadura in acetate buffered diethyl ether at 30 . The isopropylidene protective group was removed by heating the isopropylidene PtdGro diastereomers at 100 in trimethylborate in the presence of boric acid[49]. Using the method, we have found that some bacterial[47,52,53] and mammalian PtdGro molecules contain a significant amount of R,R isomer, of which existence had not been previously reported(Figure 6 C, 6 D).…”

Simple Thermodynamically-Derived Model for Predicting the Hydrolase and Transferase Activity of Phospholipase D in the Synthesis of Phosphatidylglycerol

“…where k is the rate constant, k B is the Boltzmann constant, h is the plank constant, ΔG ‡ is Gibbs free energy, and ΔH ‡ is enthalpy of activation: 32,33…”

“…where k is the rate constant, k B is the Boltzmann constant, h is the plank constant, ∆G ‡ is Gibbs free energy, and ∆H ‡ is enthalpy of activation: 32,33 Figure 4. Transphosphatidylation of phosphatidylcholine with solketal to forming phosphatidylglycerol via iso-phosphatidylglycerol.…”