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Abstract：
The human NQO1 (hNQO1) is a flavin adenine nucleotide (FAD)-dependent oxidoreductase that catalyzes the two-electron reduction of quinones to hydroquinones, being essential for the antioxidant defense system, stabilization of tumor suppressors, and activation of quinone-based chemotherapeutics. Moreover, it is overexpressed in several tumors, which makes it an attractive cancer drug target. To decipher new structural insights into the flavin reductive half-reaction of the catalytic mechanism of hNQO1, we have carried serial crystallography experiments at new ID29 beamline of the ESRF to determine, to the best of our knowledge, the first structure of the hNQO1 in complex with NADH. We have also performed molecular dynamics simulations of free hNQO1 and in complex with NADH. This is the first structural evidence that the hNQO1 functional cooperativity is driven by structural communication between the active sites through long-range propagation of cooperative effects across the hNQO1 structure. Both structural results and MD simulations have supported that the binding of NADH significantly decreases protein dynamics and stabilizes hNQO1 especially at the dimer core and interface. Altogether, these results pave the way for future time-resolved studies, both at x-ray free-electron lasers and synchrotrons, of the dynamics of hNQO1 upon binding to NADH as well as during the FAD cofactor reductive half-reaction. This knowledge will allow us to reveal unprecedented structural information of the relevance of the dynamics during the catalytic function of hNQO1.
摘要：
人类NQO1（hNQO1）是一种黄素腺嘌呤核苷酸（FAD）依赖性氧化还原酶，可催化醌的两电子还原为氢醌，对抗氧化防御系统至关重要，稳定肿瘤抑制因子，和基于醌的化疗药物的活化。此外，它在几种肿瘤中过度表达，这使得它成为一个有吸引力的抗癌药物靶点。为了破译对hNQO1催化机理的黄素还原半反应的新结构见解，我们在ESRF的新ID29光束线处进行了系列晶体学实验，以确定，据我们所知,hNQO1的第一个结构与NADH复合。我们还进行了游离hNQO1和与NADH复合的分子动力学模拟。这是第一个结构证据，表明hNQO1功能协同性是由活性位点之间的结构通讯通过跨hNQO1结构的协同作用的远距离传播来驱动的。结构结果和MD模拟都支持NADH的结合显着降低蛋白质动力学并稳定hNQO1，尤其是在二聚体核心和界面处。总之,这些结果为未来的时间分辨研究铺平了道路，在X射线自由电子激光器和同步加速器上，与NADH结合以及FAD辅因子还原半反应过程中hNQO1的动力学。这些知识将使我们能够揭示hNQO1催化功能过程中动力学相关性的前所未有的结构信息。
