Bénard Camille, Gibon Yves
UMR1332 Biologie du Fruit et Pathologie and Plateforme Métabolome, Centre de Génomique Fonctionnelle Bordeaux, INRA-Bordeaux and Bordeaux University, Villenave d'Ornon, France.
Curr Protoc Plant Biol. 2016 Mar;1(2):247-262. doi: 10.1002/cppb.20003.
The measurement of enzyme activities represents an important step towards the understanding of biological networks. Continuous or discontinuous assays can be used, as well as highly sensitive assays, depending on the abundance of the enzymes under study. To exemplify such methods, two protocols for phosphoenolpyruvate carboxylase activity (EC 4.1.1.31) in plant extracts are given. For this, an extraction protocol is also described. Then, an optimization protocol for enzyme assays using enzymatic, chemical, or biological standards is proposed. This protocol evaluates in one run the optimal extract dilution, the recovery of a standard, and the technical error in a given matrix. The interest of using biological standard in routine measurements is highlighted. © 2016 by John Wiley & Sons, Inc.
酶活性的测定是理解生物网络的重要一步。根据所研究酶的丰度，可以使用连续或不连续测定法，以及高灵敏度测定法。为举例说明此类方法，给出了两种植物提取物中磷酸烯醇式丙酮酸羧化酶活性（EC 4.1.1.31）的实验方案。为此，还描述了一种提取方案。然后，提出了一种使用酶标、化学标或生物标对酶测定进行优化的方案。该方案在一次实验中评估最佳提取物稀释度、标准品的回收率以及给定基质中的技术误差。强调了在常规测量中使用生物标的意义。© 2016约翰威立国际出版公司。
