초록
Phosphatidylserine (PS) and docosahexaenoic acid-phosphatidylserine (DHA-PS) have significant nutritional and biological functions, which are extensively used in functional food industries. Phospholipase D (PLD)-mediated transphosphatidylation of phosphatidylcholine (PC) or DHA-PC with l-serine, is an effective method for PS and DHA-PS preparation. However, because of the hydrolysis activity of PLD, PC and DHA-PC would be converted to the undesirable byproduct, phosphatidic acid (PA) and DHA-PA. In this study, a novel phospholipase D (PLD<SUB>a2</SUB>) was firstly cloned from Acinetobacter radioresistens a2 with high transphosphatidylation activity and no hydrolysis activity. In the PLD-catalyzed synthesis process (12h), both the transphosphatidylation conversion rate and selectivity of PS and DHA-PS were about 100%, which is the only PLD enzyme reported with this superiority up till now. In comparison with the majority of other known PLDs, PLD<SUB>a2</SUB> exerted the highest activity at neutral pH, and it was stable from pH 4.0 to pH 9.0. In addition, PLD<SUB>a2</SUB> had excellent thermal stability, with an optimum reaction temperature of 40<SUP>o</SUP>C and keeping more than 80% activity from 20<SUP>o</SUP>C to 60<SUP>o</SUP>C. The high catalytic selectivity mechanism of PLD<SUB>a2</SUB> was explained by utilizing homology modeling, two-step docking, and binding energy and conformation analysis. PLD<SUB>a2</SUB> ensured a stable supply of the biocatalyst with its most preponderant transphosphatidylation activity and PS selectivity, and had great potential in phospholipids industrial production.