School of Food Science and Engineering, South China University of Technology, Guangzhou, Guangdong 510640, People's Republic of China.
Guangdong Youmei Institute of Inteligent Bio-manufacturing Co., Ltd., Foshan, Guangdong 528200, People's Republic of China.
J Agric Food Chem. 2021 Sep 22;69(37):11110-11120. doi: 10.1021/acs.jafc.1c04078. Epub 2021 Sep 13.
The mechanism of active site loops of phospholipase D (PLD) binding to the lipid-water interface for catalytic reactions still remains elusive. A flexible loop (residues 376-382) in the active site of PLD (SkPLD) is conserved within PLDs in most of the species. The residue Ser380 was found to be essential for the enzyme's adsorption to the interface and its substrate recognition. The S380V mutant showed a 4.8 times higher catalytic efficiency and nearly seven times higher adsorption equilibrium coefficient compared to the wild-type SkPLD. The monolayer film technique has confirmed that the substitution of Ser380 with valine in the loop exhibited positive interaction between the enzyme and PCs with different acyl chain lengths. The results of the interfacial binding properties indicated that the S380V mutant might display suitable phosphatidylserine synthesis activity. The present study will be helpful to explain the role of residue 380 in the active site loops of PLD.
磷脂酶 D（PLD）活性位点环与脂水界面结合进行催化反应的机制仍难以捉摸。PLD（SkPLD）活性位点中的一个柔性环（残基 376-382）在大多数物种的 PLD 中是保守的。研究发现，残基 Ser380 对于酶吸附到界面及其底物识别是必需的。与野生型 SkPLD 相比，S380V 突变体的催化效率高 4.8 倍，吸附平衡常数高近 7 倍。单层膜技术证实，环中丝氨酸 380 被缬氨酸取代后，酶与具有不同酰链长度的 PCs 之间表现出正相互作用。界面结合特性的结果表明，S380V 突变体可能表现出合适的磷脂酰丝氨酸合成活性。本研究将有助于解释残基 380 在 PLD 活性位点环中的作用。
