Oda Masayuki, Xi Zhaoyong, Inaba Satomi, Slack Ryan L, Ishima Rieko
Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, 1-5 Hangi-cho, Shimogamo, Sakyo-ku, Kyoto, Kyoto 606-8522, Japan.
Department of Structural Biology, University of Pittsburgh School of, Medicine, Pittsburgh, Pennsylvania 15260, United States.
J Therm Anal Calorim. 2019 Mar;135(5):2647-2653. doi: 10.1007/s10973-018-7445-y. Epub 2018 Jun 9.
Metal-protein interactions are not necessarily tight in many transient biological processes, such as cellular signaling, enzyme regulation, and molecular recognition. Here, we analyzed the binding thermodynamics and characterized the structural effect of divalent metal ions, i.e. Mn, Zn, and Mg, to the isolated ribonuclease H (RNH) of human immunodeficiency virus (HIV) using isothermal titration calorimetry (ITC) and circular dichroism. The binding thermodynamics of Mg to RNH was determined using competition ITC experiments, and the binding affinity of Mg was found to be about 40- and 400-times lower than those of Mn and of Zn, respectively. The structural analysis showed that Mg binding had little effect on the thermal stability of RNH, while Zn and Mn binding increased the stability. The thermodynamic characteristics of RNH metal binding, compared to intact HIV reverse transcriptase, and a possible mechanism of conformational change induced upon metal ion binding, in correlation with the structure-function relationship, are discussed.
在许多瞬时生物过程中，如细胞信号传导、酶调节和分子识别，金属 - 蛋白质相互作用不一定紧密。在此，我们使用等温滴定量热法（ITC）和圆二色性分析了二价金属离子（即锰、锌和镁）与人免疫缺陷病毒（HIV）分离的核糖核酸酶H（RNH）的结合热力学，并表征了其结构效应。通过竞争ITC实验确定了镁与RNH的结合热力学，发现镁的结合亲和力分别比锰和锌低约40倍和400倍。结构分析表明，镁的结合对RNH的热稳定性影响很小，而锌和锰的结合则提高了稳定性。讨论了与完整HIV逆转录酶相比，RNH金属结合的热力学特征，以及金属离子结合诱导的构象变化的可能机制，及其与结构 - 功能关系的相关性。
