- Department of Botany, University of Delhi, Delhi 110007, India.
- Department of Biotechnology, Indian Institute of Technology, Roorkee, Uttarakhand 247667, India.
Plant Physiol Biochem. 2018 Jul;128:178-184. doi: 10.1016/j.plaphy.2018.04.039. Epub 2018 May 8.
Phosphatidic acid (PA) is an important lipid signaling molecule which interacts with Arabidopsis thaliana Sphingosine kinase1 (AtSPHK1) during several abiotic stresses particularly drought stress as a result of Abscisic acid (ABA) signaling in guard cells. PA molecules respond by generating lipid signal and/or by binding and translocating target proteins to membrane. However, site of interaction and role of PA binding to AtSPHK1 is not clear yet. Owing to the importance of AtSPHK1 during stress signaling it is imperative to decipher the site of PA interaction with AtSPHK1. To identify the PA binding region of AtSPHK1, various deletion fragments from N-terminal and C-terminal region were prepared. Results from protein lipid overlay assay using various truncated proteins of AtSPHK1 suggested the involvement of N-terminal region, between 110 and 205 amino acids, in binding with PA. In-silico analyses performed to build homologous structure of AtSPHK1 revealed that PA docking occurs in the hydrophobic cavity of DAG-Kinase domain. Deletion of amino acids VSGDGI perturbed PA-AtSPHK1 binding, indicating an essential role of these six amino acids in PA-AtSPHK1 binding.
磷酸脂酸（PA）是一种重要的脂质信号分子，在几种非生物胁迫（特别是干旱胁迫）中，它与拟南芥 Sphingosine kinase1（AtSPHK1）相互作用，这是由于脱落酸（ABA）信号在保卫细胞中的作用。PA 分子通过生成脂质信号和/或结合并将靶蛋白转运到膜上来响应。然而，PA 与 AtSPHK1 结合的作用位点尚不清楚。由于 AtSPHK1 在胁迫信号转导中的重要性，因此必须破译 PA 与 AtSPHK1 相互作用的位点。为了鉴定 AtSPHK1 的 PA 结合区域，从 N 端和 C 端区域制备了各种缺失片段。使用 AtSPHK1 的各种截断蛋白进行蛋白质脂质覆盖测定的结果表明，N 端区域（110 到 205 个氨基酸之间）参与与 PA 的结合。为构建 AtSPHK1 的同源结构而进行的计算机分析表明，PA 对接发生在 DAG-激酶结构域的疏水区。氨基酸 VSGDGI 的缺失破坏了 PA-AtSPHK1 的结合，表明这六个氨基酸在 PA-AtSPHK1 结合中起着重要作用。
