Cooney Michael J
Hawaii Natural Energy Institute, School of Ocean and Earth Science and Technology, University of Hawaii at Manoa, 1680 East-west Rd., Honolulu, HI, 96822, USA.
Methods Mol Biol. 2017;1504:215-232. doi: 10.1007/978-1-4939-6499-4_17.
Enzyme kinetics is the study of the chemical reactions that are catalyzed by enzymes, with a focus on their reaction rates. The study of an enzyme's kinetics considers the various stages of activity, reveals the catalytic mechanism of this enzyme, correlates its value to assay conditions, and describes how a drug or a poison might inhibit the enzyme. Victor Henri initially reported that enzyme reactions were initiated by a bond between the enzyme and the substrate. By 1910, Michaelis and Menten were advancing their work by studying the kinetics of an enzyme saccharase which catalyzes the hydrolysis of sucrose into glucose and fructose. They published their analysis and ever since the Michaelis-Menten equation has been used as the standard to describe the kinetics of many enzymes. Unfortunately, soluble enzymes must generally be immobilized to be reused for long times in industrial reactors. In addition, other critical enzyme properties have to be improved like stability, activity, inhibition by reaction products, and selectivity towards nonnatural substrates. Immobilization is by far the chosen process to achieve these goals.Although the Michaelis-Menten approach has been regularly adapted to the analysis of immobilized enzyme activity, its applicability to the immobilized state is limited by the barriers the immobilization matrix places upon the measurement of compounds that are used to model enzyme kinetics. That being said, the estimated value of the Michaelis-Menten coefficients (e.g., V , K ) can be used to evaluate effects of immobilization on enzyme activity in the immobilized state when applied in a controlled manner. In this review enzyme activity and kinetics are discussed in the context of the immobilized state, and a few novel protocols are presented that address some of the unique constraints imposed by the immobilization barrier.
酶动力学是对由酶催化的化学反应的研究，重点是它们的反应速率。对酶动力学的研究考虑了活性的各个阶段，揭示了这种酶的催化机制，将其值与测定条件相关联，并描述了药物或毒物如何抑制该酶。维克多·亨利最初报道酶反应是由酶与底物之间的键引发的。到1910年，米氏（Michaelis）和门滕（Menten）通过研究催化蔗糖水解为葡萄糖和果糖的蔗糖酶的动力学来推进他们的工作。他们发表了他们的分析，从那时起，米氏方程就一直被用作描述许多酶动力学的标准。不幸的是，可溶性酶通常必须固定化才能在工业反应器中长期重复使用。此外，还必须改善其他关键的酶特性，如稳定性、活性、被反应产物抑制以及对非天然底物的选择性。固定化是迄今为止实现这些目标所选择的方法。尽管米氏方法已经常被用于分析固定化酶的活性，但其在固定化状态下的适用性受到固定化基质对用于模拟酶动力学的化合物测量所造成的障碍的限制。话虽如此，当以可控方式应用时，米氏系数（例如V、K）的估计值可用于评估固定化对固定化状态下酶活性的影响。在这篇综述中，将在固定化状态的背景下讨论酶活性和动力学，并提出一些新颖的方案，以解决固定化障碍所带来的一些独特限制。
