Measurement of Enzyme Activities and Optimization of Continuous and Discontinuous Assays

Q1 Agricultural and Biological Sciences Current protocols in plant biology Pub Date : 2016-06-10 DOI:10.1002/cppb.20003

Camille Bénard, Yves Gibon

引用次数: 7

Abstract

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.

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酶活性的测定及连续和间断试验的优化

酶活性的测量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，以及高灵敏度测定。为了举例说明这些方法，给出了植物提取物中磷酸烯醇丙酮酸羧化酶活性(EC 4.1.1.31)的两种方案。为此，还描述了一种提取协议。然后，提出了酶、化学或生物标准酶分析的优化方案。本方案在一次运行中评估最佳萃取物稀释度、标准品回收率和给定基质中的技术误差。强调了在常规测量中使用生物标准的重要性。©2016 by John Wiley &儿子,Inc。

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Current protocols in plant biology Agricultural and Biological Sciences-Plant Science

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期刊介绍： Sound and reproducible laboratory methods are the foundation of scientific discovery. Yet nuances that are critical for an experiment''s success are not captured in the primary literature but exist only as part of a lab''s oral tradition. Current Protocols in Plant Biology provides reproducible step-by-step instructions for protocols relevant to plant research. Furthermore, Current Protocols content is thoughtfully organized by topic for optimal usage and to maximize contextual knowledge. Quarterly issues allow Current Protocols in Plant Biology to constantly evolve to keep pace with the newest discoveries and developments. Current Protocols in Plant Biology is the comprehensive source for protocols in the multidisciplinary field of plant biology, providing an extensive range of protocols from basic to cutting edge. Coverage includes: Extraction and analysis of DNA, RNA, proteins Chromosome analysis Transcriptional analysis Protein expression Metabolites Plant enzymology Epigenetics Plant genetic transformation Mutagenesis Arabidopsis, Maize, Poplar, Rice, and Soybean, and more.