一种针对SIRT1、SIRT2和SIRT3的改进型荧光检测法。
An improved fluorogenic assay for SIRT1, SIRT2, and SIRT3.
作者信息
Chiang Ying-Ling, Lin Hening
机构信息
Department of Chemistry and Chemical Biology, Baker Laboratory, Cornell University, Ithaca, NY 14853, USA.
出版信息
Org Biomol Chem. 2016 Feb 21;14(7):2186-90. doi: 10.1039/c5ob02609a. Epub 2016 Jan 21.
Sirtuins are NAD-dependent lysine deacylases that play critical roles in cellular regulation and are implicated in human diseases. Modulators of sirtuins are needed as tools for investigating their biological functions and possible therapeutic applications. However, the discovery of sirtuin modulators is hampered by the lack of efficient sirtuin assays. Here we report an improved fluorogenic assay for SIRT1, SIRT2, and SIRT3 using a new substrate, a myristoyl peptide with a C-terminal aminocoumarin. The new assay has several advantages, including significantly lower substrate concentration needed, increased signal-to-background ratio, and improved Z'-factor. The novel assay thus will expedite high-throughput screening of SIRT1, SIRT2, and SIRT3 modulators.
沉默调节蛋白是依赖烟酰胺腺嘌呤二核苷酸（NAD）的赖氨酸脱酰基酶，在细胞调节中起关键作用，并与人类疾病有关。需要沉默调节蛋白的调节剂作为研究其生物学功能和可能的治疗应用的工具。然而，由于缺乏有效的沉默调节蛋白检测方法，沉默调节蛋白调节剂的发现受到阻碍。在此，我们报告了一种针对沉默信息调节因子1（SIRT1）、沉默信息调节因子2（SIRT2）和沉默信息调节因子3（SIRT3）的改进的荧光检测方法，该方法使用一种新的底物，即一种带有C端氨基香豆素的肉豆蔻酰肽。这种新的检测方法有几个优点，包括所需底物浓度显著降低、信号背景比增加和Z'因子改善。因此，这种新的检测方法将加快对SIRT1、SIRT2和SIRT3调节剂的高通量筛选。
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