一种基于发光的实验方案，用于通过定量小鼠或人肝细胞中的酮己糖激酶酶活性来评估果糖代谢。
A luminescence-based protocol for assessing fructose metabolism via quantification of ketohexokinase enzymatic activity in mouse or human hepatocytes.
作者信息
Park Se-Hyung, Helsley Robert N, Noetzli Leila, Tu Ho-Chou, Wallenius Kristina, O'Mahony Gavin, Boucher Jeremie, Liu Jianming, Softic Samir
机构信息
Department of Pediatrics, Division of Pediatric Gastroenterology, University of Kentucky College of Medicine, 900 South Limestone, Wethington Rm 527, Lexington, KY 40536, USA.
Alnylam Pharmaceuticals Inc., Cambridge, MA 021428, USA.
出版信息
STAR Protoc. 2021 Aug 9;2(3):100731. doi: 10.1016/j.xpro.2021.100731. eCollection 2021 Sep 17.
Ketohexokinase (KHK) catalyzes the first step of fructose metabolism. Inhibitors of KHK enzymatic activity are being evaluated in clinical trials for the treatment of non-alcoholic fatty liver disease (NAFLD) and diabetes. Here, we present a luminescence-based protocol to quantify KHK activity. The accuracy of this technique has been validated using knockdown and overexpression of KHK and . The specificity of the assay has been verified using 3-O-methyl-D-fructose, a non-metabolizable analog of fructose, heat inactivation of hexokinases, and depletion of potassium. For complete details on the use of this protocol, please refer to Damen et al. (2021).
酮己糖激酶（KHK）催化果糖代谢的第一步。KHK酶活性抑制剂正在进行治疗非酒精性脂肪性肝病（NAFLD）和糖尿病的临床试验评估。在此，我们提出一种基于发光的方法来定量KHK活性。该技术的准确性已通过KHK的敲低和过表达得到验证。该检测方法的特异性已通过3-O-甲基-D-果糖（一种果糖的非代谢类似物）、己糖激酶的热失活和钾离子耗尽得到验证。有关该方法使用的完整详细信息，请参考达门等人（2021年）的文献。
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