Wang Lei, Li Zihuan, Xuan Yumi, Qin Jingkun, Li Shuju, Zhong Fumei, Song Yuexiao, Yang Kanglong, Lv Mengqi, Li Fudong, Jiahai Zhang, Pan Yueyin, Guang Shouhong, Zhao Yuzheng, Shi Yunyu, Liu Xing, Du Yingying, Gao Jia, Ruan Ke
Department of Oncology, the First Affiliated Hospital & School of Life Sciences, Ministry of Education Key Laboratory for Membrane-less Organelles & Cellular Dynamics, Hefei National Research Center for Interdisciplinary Sciences at the Microscale, Biomedical Sciences and Health Laboratory of Anhui Province, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, 230027, China.
Department of Oncology, the First Affiliated Hospital of Anhui Medical University, Hefei, 230022, China.
Nat Commun. 2025 Jul 25;16(1):6859. doi: 10.1038/s41467-025-61732-y.
Human guanylate kinase (GMPK) as the sole enzyme for GDP biosynthesis plays pivotal roles in antiviral prodrug activation and tumorigenesis. Despite its biological significance, the catalytic mechanism remains poorly understood. Here, we resolve crystal structures of GMPK in free and GMP-bound form, revealing the interdomain motions of GMPBD and LID relative to the CORE domain. Biochemical assays demonstrate potassium's dual functionality in substrate recognition and phosphoryl transfer catalysis. Structural analyses uncover intradomain conformational motion within the LID domain and essential interactions for ADP/ATP binding. Notably, the cooperative ATPγS binding potentiated by prior GMP binding are structurally elucidated. Three key complexes, pre-reaction state (GMP/ATPγS), transition state (AlF mimic), and post-reaction state (GDP/ADP), collectively delineate the reversible catalytic pathway. This comprehensive structural characterization of GMPK's dynamic landscape establishes a foundation for developing conformation-specific inhibitors through structure-guided drug design.
人鸟苷酸激酶（GMPK）作为GDP生物合成的唯一酶，在抗病毒前药激活和肿瘤发生中起关键作用。尽管其具有生物学意义，但其催化机制仍知之甚少。在这里，我们解析了游离形式和GMP结合形式的GMPK晶体结构，揭示了GMPBD和LID相对于CORE结构域的结构域间运动。生化分析证明了钾在底物识别和磷酸转移催化中的双重功能。结构分析揭示了LID结构域内的结构域内构象运动以及ADP/ATP结合的关键相互作用。值得注意的是，在结构上阐明了先前GMP结合增强的协同ATPγS结合。三种关键复合物，即反应前状态（GMP/ATPγS）、过渡状态（AlF模拟物）和反应后状态（GDP/ADP），共同描绘了可逆催化途径。对GMPK动态景观的这种全面结构表征为通过结构导向药物设计开发构象特异性抑制剂奠定了基础。
