Maliga Zoltan, Xing Jun, Cheung Herbert, Juszczak Laura J, Friedman Joel M, Rosenfeld Steven S
Program in Biophysics, Harvard University, Boston, Massachusetts 02115, USA.
J Biol Chem. 2006 Mar 24;281(12):7977-82. doi: 10.1074/jbc.M511955200. Epub 2006 Jan 23.
Monastrol is a small molecule inhibitor that is specific for Eg5, a member of the kinesin 5 family of mitotic motors. Crystallographic models of Eg5 in the presence and absence of monastrol revealed that drug binding produces a variety of structural changes in the motor, including in loop L5 and the neck linker. What is not clear from static crystallographic models, however, is the sequence of structural changes produced by drug binding. Furthermore, because crystallographic structures can be influenced by the packing forces in the crystal, it also remains unclear whether these drug-induced changes occur in solution, at physiologically active concentrations of monastrol or of other drugs that target this site. We have addressed these issues by using a series of spectroscopic probes to monitor the structural consequences of drug binding. Our results demonstrated that the crystallographic model of an Eg5-ADP-monastrol ternary complex is consistent with several solution-based spectroscopic probes. Furthermore, the kinetics of these spectroscopic signal changes allowed us to determine the temporal sequence of drug-induced structural transitions. These results suggested that L5 may be an element in the pathway that links the state of the nucleotide-binding site to the neck linker in kinesin motors.
莫那可尔是一种小分子抑制剂，对驱动蛋白5家族有丝分裂马达成员Eg5具有特异性。在有和没有莫那可尔的情况下，Eg5的晶体学模型表明，药物结合会在马达中产生多种结构变化，包括在环L5和颈部连接区。然而，从静态晶体学模型中尚不清楚药物结合产生的结构变化顺序。此外，由于晶体学结构可能会受到晶体中堆积力的影响，因此也不清楚这些药物诱导的变化是否在溶液中、在生理活性浓度的莫那可尔或其他靶向该位点的药物作用下发生。我们通过使用一系列光谱探针来监测药物结合的结构后果，解决了这些问题。我们的结果表明，Eg5-ADP-莫那可尔三元复合物的晶体学模型与几种基于溶液的光谱探针一致。此外，这些光谱信号变化的动力学使我们能够确定药物诱导的结构转变的时间顺序。这些结果表明，L5可能是在驱动蛋白马达中将核苷酸结合位点的状态与颈部连接区联系起来的途径中的一个元件。
