Wang Huanchen, Liu Yudong, Huai Qing, Cai Jiwen, Zoraghi Roya, Francis Sharron H, Corbin Jackie D, Robinson Howard, Xin Zhongcheng, Lin Guiting, Ke Hengming
Department of Biochemistry and Biophysics and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina 27599-7260.
Department of Biochemistry and Biophysics and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina 27599-7260; School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, 510080, China.
J Biol Chem. 2006 Jul 28;281(30):21469-21479. doi: 10.1074/jbc.M512527200. Epub 2006 May 30.
Phosphodiesterase-5 (PDE5) is the target for sildenafil, vardenafil, and tadalafil, which are drugs for treatment of erectile dysfunction and pulmonary hypertension. We report here the crystal structures of a fully active catalytic domain of unliganded PDE5A1 and its complexes with sildenafil or icarisid II. These structures together with the PDE5A1-isobutyl-1-methylxanthine complex show that the H-loop (residues 660-683) at the active site of PDE5A1 has four different conformations and migrates 7-35A upon inhibitor binding. In addition, the conformation of sildenafil reported herein differs significantly from those in the previous structures of chimerically hybridized or almost inactive PDE5. Mutagenesis and kinetic analyses confirm that the H-loop is particularly important for substrate recognition and that invariant Gly(659), which immediately precedes the H-loop, is critical for optimal substrate affinity and catalytic activity.
磷酸二酯酶-5（PDE5）是西地那非、伐地那非和他达拉非的作用靶点，这些药物用于治疗勃起功能障碍和肺动脉高压。我们在此报告未结合配体的PDE5A1的完全活性催化结构域及其与西地那非或淫羊藿苷II复合物的晶体结构。这些结构与PDE5A1-异丁基-1-甲基黄嘌呤复合物一起表明，PDE5A1活性位点处的H环（第660-683位氨基酸残基）有四种不同构象，在抑制剂结合时迁移7-35埃。此外，本文报道的西地那非构象与先前嵌合杂交或几乎无活性的PDE5结构中的构象有显著差异。诱变和动力学分析证实，H环对底物识别尤为重要，紧接在H环之前的不变甘氨酸（Gly659）对最佳底物亲和力和催化活性至关重要。
