In-Situ Co-Cross-Linking of Phospholipase D and Poly-L-Lysine for the Highly Efficient Synthesis of Phosphatidylglycerol

Phospholipase D (PLD) was effectively immobilized on a nanocomposite support composed of ZnO nanowires and SiO 2 through in-situ co-cross-linking with poly-L-lysine (PLL). An anionic and long-chained bisepoxide cross-linker was electrostatically adsorbed on the surface of ZnO nanowires prior to co-cross-linking. Under the optimal pH, 6.5, the loading amount of PLD reached an average of 114 mg per g support for PLL concentrations in the range of 0–0.6 mg mL −1 . Co-cross-linking of PLD with PLL enhanced the activity of PLD remarkably. The highest specific activity of co-cross-linked PLD reached 25.8 U per mg protein , which was 1.62 times as high as that of the cross-linked PLD (15.9 U per mg protein ). (One unit of PLD activity [U] refers to the amount of PLD, which liberates 1 μmol of p-nitrophenol per min via hydrolysis of phosphatidyl-p-nitrophenol at pH 8.0 and 30 °C). Under the optimized conditions, the yield of phosphatidylglycerol (PtdGro) reached 98.6% within 80 min at 45 °C and pH 6.5. The co-cross-linked PLD not only exhibited good resistance to pH and heat inactivation but also improved storage stability and reusability. The yield of PtdGro remained ≥85.4% after 13 repeat uses.