Department of Physiology and Biophysics and The Massey Cancer Center, Virginia Commonwealth University School of Medicine, Richmond, VA 23298, United States of America.
Department of Physiology and Biophysics and The Massey Cancer Center, Virginia Commonwealth University School of Medicine, Richmond, VA 23298, United States of America.
J Inorg Biochem. 2020 Aug;209:111120. doi: 10.1016/j.jinorgbio.2020.111120. Epub 2020 May 21.
Cytochromes P450 (CYPs) display remarkable plasticity in their ability to bind substrates and catalyze a broad array of chemical reactions. Herein we evaluate binding of androstenedione, testosterone, and 7-hydroxyflavone to CYP19A1, also known as aromatase, in phospholipid nanodiscs by stopped-flow UV-vis spectroscopy. Exponential fitting of the kinetic traces supports the possibility of a multi-step binding mechanism. Subsequent global fitting of the data to the solutions of the coupled differential equations describing the fundamental mechanisms of induced fit and conformational selection, consistently support presence of the latter. To our knowledge, this is the first discrimination of conformational selection from induced fit for a mono-disperse CYP in a native-like membrane environment. In addition, 7-hydroxyflavone binds to CYP19A1 nanodiscs with comparable affinity to the substrates and induces an unusual spectral response likely attributable to hydrogen bonding to, rather than displacement of the heme-coordinated water molecule.
细胞色素 P450（CYPs）在结合底物和催化广泛的化学反应方面表现出显著的可塑性。在此，我们通过停流紫外可见光谱法评估了雄烯二酮、睾酮和 7-羟基黄酮与磷脂纳米盘中的 CYP19A1（也称为芳香酶）的结合。动力学轨迹的指数拟合支持多步结合机制的可能性。随后，将数据全局拟合到描述诱导契合和构象选择基本机制的耦合微分方程的解中，一致支持后一种机制的存在。据我们所知，这是在天然样膜环境中单分散 CYP 中首次从诱导契合中区分构象选择。此外，7-羟基黄酮与 CYP19A1 纳米盘的结合亲和力与底物相当，并诱导出一种不寻常的光谱响应，可能归因于与氢键的相互作用，而不是取代血红素配位的水分子。
