Department of Cell and Molecular Biology, Uppsala University, Husarg. 3, BMC Box 596, SE-75124 Uppsala, Sweden.
Biochimie. 2011 Sep;93(9):1623-9. doi: 10.1016/j.biochi.2011.05.031. Epub 2011 Jun 12.
Enzyme inhibitors are used in many areas of the life sciences, ranging from basic research to the combat of disease in the clinic. Inhibitors are traditionally characterized by how they affect the steady-state kinetics of enzymes, commonly analyzed on the assumption that enzyme-bound and free substrate molecules are in equilibrium. This assumption, implying that an enzyme-bound substrate molecule has near zero probability to form a product rather than dissociate, is valid only for very inefficient enzymes. When it is relaxed, more complex but also more information-rich steady-state kinetics emerges. Although solutions to the general steady-state kinetics problem exist, they are opaque and have been of limited help to experimentalists. Here we reformulate the steady-state kinetics of enzyme inhibition in terms of new parameters. These allow for assessment of ambiguities of interpretation due to kinetic scheme degeneracy and provide an intuitively simple way to analyze experimental data. We illustrate the method by concrete examples of how to assess scheme degeneracy and obtain experimental estimates of all available rate and equilibrium constants. We suggest simple, complementary experiments that can remove ambiguities and greatly enhance the accuracy of parameter estimation.
酶抑制剂在生命科学的许多领域都有应用，从基础研究到临床疾病的治疗。抑制剂通常根据它们如何影响酶的稳态动力学来进行特征描述，通常的分析假设是酶结合和游离的底物分子处于平衡状态。这个假设意味着酶结合的底物分子形成产物的概率几乎为零，而不是解离，这个假设仅适用于效率非常低的酶。当这个假设放松时，会出现更复杂但也更丰富信息的稳态动力学。尽管存在一般稳态动力学问题的解决方案，但它们不透明，对实验人员的帮助有限。在这里，我们根据新的参数重新表述酶抑制的稳态动力学。这些参数允许评估由于动力学方案简并性而导致的解释歧义，并提供了一种直观简单的方法来分析实验数据。我们通过具体的例子来说明如何评估方案简并性，并获得所有可用速率和平衡常数的实验估计值。我们建议进行简单、互补的实验，可以消除歧义并大大提高参数估计的准确性。
