Quantitative Analysis of Polyphosphoinositide, Bis(monoacylglycero)phosphate, and Phosphatidylglycerol Species by Shotgun Lipidomics After Methylation.
Quantitative Analysis of Polyphosphoinositide, Bis(monoacylglycero)phosphate, and Phosphatidylglycerol Species by Shotgun Lipidomics After Methylation.
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DOI：
10.1007/978-1-0716-1410-5_6
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Han X
中科院分区：
文献类型：
--
作者：
Pan M;Qin C;Han X
磷脂在生物过程中发挥着重要的作用，即使在很低的水平上也是如此。例如，双(单甘油)磷酸(BMP)参与了溶酶体贮藏疾病的发病过程，聚磷脂酰肌醇(PPI)在细胞信号和功能中发挥着关键作用。磷脂酰甘油(PG)是骨形态发生蛋白(BMP)的结构异构体，介导脂-蛋白和脂-脂相互作用，抑制血小板活化因子和磷脂酰胆碱的转运。然而，由于这些磷脂的丰度较低，从生物样本中分析这些磷脂在技术上具有挑战性。因此，这些磷脂的细胞功能和代谢仍然难以捉摸。本章综述了三甲基硅基重氮甲烷(TMS-D)甲基化后的一种新的鸟枪脂组学方法，用于准确和全面地分析生物样品中的这些磷脂物种。首先，采用改进的Bligh和Dyer方法提取组织脂用于PPI分析，而采用改进的甲基叔丁基醚(MTBE)提取法和改进的Folch提取法提取组织脂用于PPI分析。其次，分别对PG/BMP和PPI进行TMS-D甲基化。然后，我们描述了可以作为经济有效和相对高通量的方法来确定骨形态发生蛋白、PG和PPI物种及其不同磷酸盐位置异构体(S)和脂肪酰链的鸟枪式脂肪组学策略。甲基化后的鸟枪脂组学方法实现了对低丰度脂类的可行和可靠的定量分析。这一新方法的应用将使我们能够揭示这些磷脂在健康和疾病状态下的代谢和功能。
Phospholipids play important roles in biological process even at a very low level. For example, bis(monoacylglycerol)phosphate (BMP) is involved in the pathogenesis of lysosomal storage diseases, polyphosphoinositides (PPI) play critical roles in cellular signaling and functions. Phosphatidylglycerol (PG), a structural isomer of BMP, mediates lipid-protein and lipid-lipid interactions, and inhibits platelet activating factor and phosphatidylcholine transferring. However, due to their low abundance, the analysis of these phospholipids from biological samples is technically challenging. Therefore, the cellular function and metabolism of these phospholipids are still elusive. This chapter overviews a novel method of shotgun lipidomics after methylation with trimethylsilyl-diazomethane (TMS-D) for accurate and comprehensive analysis of these phospholipid species in biological samples. Firstly, a modified Bligh and Dyer procedure is performed to extract tissue lipids for PPI analysis, whereas modified methyl-tert-butylether (MTBE) extraction and modified Folch extraction methods are described to extract tissue lipids for PPI analysis. Secondly, TMS-D methylation is performed to derivatize PG/BMP and PPI, respectively. Then, we described the shotgun lipidomics strategies that can be used as cost-effective and relatively high throughput methods to determine BMP, PG, and PPI species and isomers with different phosphate position(s) and fatty acyl chains. The described method of shotgun lipidomics after methylation achieves feasible and reliable quantitative analysis of low-abundance lipid classes. The application of this novel method should enable us to reveal the metabolism and functions of these phospholipids in healthy and disease states.