Department of Biology, Appalachian State University, 572 Rivers Street, Boone, NC 28608-2027, USA.
J Phycol. 2012 Jun;48(3):580-4. doi: 10.1111/j.1529-8817.2012.01137.x. Epub 2012 Apr 24.
Common methods for assaying acyl-CoA:diacylglycerol acyltransferase (DGAT) enzymatic activity rely upon radiolabeled substrates or product assay. We developed a novel assay that directly quantifies endogenous DGAT activity through the use of a fluorescently labeled substrate. We performed this assay with microsomal protein, 2-(6-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoyl-1-hexadecanoyl-sn-glycero-3-diacylglycerol (NBD-DAG), and oleoyl-CoA substrates. DGAT activity was analyzed in three species of algae as well as rat liver. The protocol proved to be sensitive and reliable. This assay may be used to facilitate research in the areas of biodiesel, oilseed crops, and triacylglycerol-related human pathologies.
常用的酰基辅酶 A：二酰基甘油酰基转移酶（DGAT）酶活性测定方法依赖于放射性标记的底物或产物测定。我们开发了一种新的测定方法，通过使用荧光标记的底物直接定量内源性 DGAT 活性。我们使用微粒体蛋白、2-（6-（7-硝基苯并-2-氧代-1,3-二唑-4-基）氨基）己酰基-1-十六烷酰基-sn-甘油-3-二酰基甘油（NBD-DAG）和油酰辅酶 A 底物进行了这项测定。在三种藻类以及大鼠肝脏中分析了 DGAT 活性。该方案被证明是敏感和可靠的。该测定法可用于促进生物柴油、油籽作物和与三酰基甘油相关的人类病理学领域的研究。
