Department of Chemistry, Louisiana State University, 232 Choppin Hall, Baton Rouge, Louisiana 70803-1804, USA.
Biochemistry. 2012 Oct 9;51(40):8014-26. doi: 10.1021/bi300760u. Epub 2012 Sep 28.
NAD(P)H:quinone oxidoreductase type I (NQO1) is a target enzyme for triggered delivery of drugs at inflamed tissue and tumor sites, particularly those that challenge traditional therapies. Prodrugs, macromolecules, and molecular assemblies possessing trigger groups that can be cleaved by environmental stimuli are vehicles with the potential to yield active drug only at prescribed sites. Furthermore, quinone propionic acids (QPAs) covalently attached to prodrugs or liposome surfaces can be removed by application of a reductive trigger stimulus, such as that from NQO1; their rates of reductive activation should be tunable via QPA structure. We explored in detail the recombinant human NAD(P)H:quinone oxidoreductase type I (rhNQO1)-catalyzed NADH reduction of a family of substituted QPAs and obtained high precision kinetic parameters. It is found that small changes in QPA structure-in particular, single atom and function group substitutions on the quinone ring at R(1)-lead to significant impacts on the Michaelis constant (K(m)), maximum velocity (V(max)), catalytic constant (k(cat)), and catalytic efficiency (k(cat)/K(m)). Molecular docking simulations demonstrate that alterations in QPA structure result in large changes in QPA alignment and placement with respect to the flavin isoalloxazine ring in the active site of rhNQO1; a qualitative relationship exists between the kinetic parameters and the depth of QPA penetration into the rhNQO1 active site. From a quantitative perspective, a very good correlation is observed between log(k(cat)/K(m)) and the molecular-docking-derived distance between the flavin hydride donor site and quinone hydride acceptor site in the QPAs, an observation that is in agreement with developing theories. The comprehensive kinetic and molecular modeling knowledge obtained for the interaction of recombinant human NQO1 with the quinone propionic acid analogues provides insight into the design and implementation of the QPA trigger groups for drug delivery applications.
NAD(P)H：醌氧化还原酶 1 型（NQO1）是一种在炎症组织和肿瘤部位靶向递药的靶标酶，尤其是那些对传统治疗方法具有挑战性的药物。前药、大分子和具有可以被环境刺激物切割的触发基团的分子组装体是具有仅在预定部位产生活性药物的潜力的载体。此外，共价连接到前药或脂质体表面的醌丙酸（QPAs）可以通过应用还原性触发刺激物（例如 NQO1）去除；它们的还原激活率可以通过 QPA 结构进行调节。我们详细研究了重组人 NAD(P)H：醌氧化还原酶 1 型（rhNQO1）催化的一系列取代 QPAs 的 NADH 还原，获得了高精度的动力学参数。结果发现，QPAs 结构的微小变化——尤其是醌环上 R（1）处的单个原子和官能团取代——对米氏常数（K(m)）、最大速度（V(max)）、催化常数（k(cat)）和催化效率（k(cat)/K(m)）有显著影响。分子对接模拟表明，QPAs 结构的改变导致 QPA 在 rhNQO1 活性部位黄素异咯嗪环中的排列和位置发生很大变化；动力学参数与 QPA 穿透 rhNQO1 活性部位的深度之间存在定性关系。从定量角度来看，观察到 k(cat)/K(m)的对数与 QPAs 中黄素氢供体位点与醌氢受体位点之间的分子对接衍生距离之间存在非常好的相关性，这一观察结果与发展理论一致。获得的关于重组人 NQO1 与醌丙酸类似物相互作用的综合动力学和分子建模知识为设计和实施用于药物输送的 QPA 触发基团提供了深入的见解。
