Okada Mayu, Tateishi Yutaka, Nojiri Eri, Mikawa Tsutomu, Rajesh Sundaresan, Ogasa Hiroki, Ueda Takumi, Yagi Hiromasa, Kohno Toshiyuki, Kigawa Takanori, Shimada Ichio, Güntert Peter, Ito Yutaka, Ikeya Teppei
Department of Chemistry, Graduate School of Science, Tokyo Metropolitan University, 1-1 minamiosawa, Hachioji, Tokyo 192-0397, Japan.
International Graduate School of Arts and Sciences, Yokohama City University, 1-7-29 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan.
J Am Chem Soc. 2025 Aug 20;147(33):29884-29894. doi: 10.1021/jacs.5c06502. Epub 2025 Aug 6.
Despite accumulating evidence that protein dynamics is indispensable for understanding the structural basis of biological activities, it remains challenging to visualize the spatial description of the dynamics and to associate transient conformations with their molecular functions. We have developed a new NMR protein structure determination method for the inference of multistate conformations using multiple types of NMR data, including paramagnetic NMR and residual dipolar couplings, as well as conventional NOEs. Integration of these data in the structure calculation permits delineating accurate ensemble structures of biomacromolecules. Applying the method to yeast ubiquitin hydrolase 1, we find large dynamics of its N-terminus (gating lid) and crossover loop surrounding the active site for ubiquitin-recognition and proteolysis. The N-terminus (gating lid) moves into and out of the crossover loop, suggesting their underlying functional significance. Our results, including those from biochemical analysis, show that large motion surrounding the active site contributes strongly to the efficiency of the enzymatic activity.
尽管越来越多的证据表明蛋白质动力学对于理解生物活性的结构基础不可或缺，但要可视化动力学的空间描述并将瞬态构象与其分子功能联系起来仍然具有挑战性。我们开发了一种新的核磁共振蛋白质结构测定方法，用于使用多种类型的核磁共振数据（包括顺磁核磁共振和剩余偶极耦合）以及传统的核Overhauser效应（NOE）来推断多态构象。在结构计算中整合这些数据可以描绘出生物大分子准确的集合结构。将该方法应用于酵母泛素水解酶1，我们发现其N端（门控盖）和围绕泛素识别与蛋白水解活性位点的交叉环存在较大的动力学变化。N端（门控盖）移入和移出交叉环，表明它们潜在的功能意义。我们的结果，包括来自生化分析的结果，表明活性位点周围的大幅度运动对酶活性的效率有很大贡献。
