使用亲水作用液相色谱-质谱法测定细胞和高尔基体组分中的3'-磷酸腺苷-5'-磷酸硫酸酯
Determination of 3'-phosphoadenosine-5'-phosphosulfate in cells and Golgi fractions using hydrophilic interaction liquid chromatography-mass spectrometry.
作者信息
Dowood Rua Kareem, Adusumalli Ravi, Tykesson Emil, Johnsen Elin, Lundanes Elsa, Prydz Kristian, Wilson Steven Ray
机构信息
Department of Chemistry, University of Oslo, Post Box 1033, Blindern, NO-0315 Oslo, Norway.
Department of Biosciences, University of Oslo, Post Box 1066, Blindern, NO-0316 Oslo, Norway.
出版信息
J Chromatogr A. 2016 Oct 28;1470:70-75. doi: 10.1016/j.chroma.2016.10.001. Epub 2016 Oct 4.
3'-Phosphoadenosine-5'-phosphosulfate (PAPS) is a key player in the sulfation of biomolecules, but methods for selective measurements are lacking. A liquid chromatography-mass spectrometry (LC-MS) approach for measuring PAPS was developed. A central feature of the method was employing hydrophilic interaction liquid chromatography (HILIC), which is highly suited for separating very polar/charged compounds, and is compatible with electrospray MS. Using simple instrumentation, the analysis time per sample was below 10min and the method was characterized by easy sample preparation. The method was used to monitor decreasing levels of PAPS as function of sodium chlorate treatment (an inhibitor of PAPS synthesis) in whole-cell lysates as well as Golgi-fractions. The method allowed PAPS to be chromatographically separated from ADP and ATP, which can interfere with measurements if a less resolving LC-MS method is used.
3'-磷酸腺苷-5'-磷酸硫酸酯（PAPS）是生物分子硫酸化过程中的关键物质，但缺乏选择性测量方法。本文开发了一种用于测量PAPS的液相色谱-质谱联用（LC-MS）方法。该方法的核心特点是采用亲水作用液相色谱（HILIC），它非常适合分离极性很强/带电荷的化合物，并且与电喷雾质谱兼容。使用简单的仪器，每个样品的分析时间不到10分钟，且该方法的特点是样品制备简便。该方法用于监测全细胞裂解物以及高尔基体组分中PAPS水平随氯酸钠处理（PAPS合成抑制剂）的降低情况。该方法能够将PAPS与ADP和ATP进行色谱分离，如果使用分辨率较低的LC-MS方法，ADP和ATP会干扰测量。
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