Wright Louwrance P, Phillips Michael A
Department of Biochemistry, Max Planck Institute for Chemical Ecology, Hans-Knöll-Str. 8, 07745, Jena, Germany,
Methods Mol Biol. 2014;1153:9-20. doi: 10.1007/978-1-4939-0606-2_2.
The first enzyme in the methylerythritol phosphate (MEP) pathway is 1-deoxy-D-xylulose 5-phosphate (DXP) synthase (DXS). As such this enzyme is considered to be important in the control of plastidial isoprenoid production. Measuring the activity of DXS in plant extracts is therefore crucial to understanding the regulation of the MEP pathway. Due to the relatively low amounts of DXS, the activity of this enzyme can only be measured using highly sensitive analytical equipment. Here, a method is described to determine the DXS enzyme activity in a crude plant extract, by measuring DXP production directly using high performance liquid chromatography linked to a tandem triple quadrupole mass spectrometry detector (LC-MS/MS).
甲基赤藓糖醇磷酸（MEP）途径中的首个酶是1-脱氧-D-木酮糖5-磷酸（DXP）合酶（DXS）。因此，该酶被认为在质体类异戊二烯生成的调控中起着重要作用。所以，测定植物提取物中DXS的活性对于理解MEP途径的调控至关重要。由于DXS的含量相对较低，只能使用高灵敏度分析设备来测定该酶的活性。在此，描述了一种通过使用与串联三重四极杆质谱检测器联用的高效液相色谱法（LC-MS/MS）直接测定DXP生成量，来确定粗植物提取物中DXS酶活性的方法。
