CRYSTAL STRUCT. OF CYCLIC NUCLEOTIDE PHOSPHODIESTERASE

描述：（逐字来自申请人的摘要） 磷酸二酯酶（PDE）催化腺苷3 '，5'-环 环磷酸鸟苷（cGMP）和环磷酸鸟苷（cAMP）， 分别产生5 ′-AMP和5 ′-GMP。PDE是一种关键酶， 细胞内的cAMP浓度被称为“第二信使”， 介导细胞对多种激素的反应， 神经传递素已对人类PDE的10个家族和22个亚型进行了研究。 鉴定22种亚型PDE的mRNA被进一步剪接以产生 超过60种PDE亚型。PDE的不同亚型位于不同的 细胞区室，并具有不同的底物特异性。这两 PDE的特性引起了制药公司的极大关注， 过去十年许多选择性PDE抑制剂已被研究用于治疗 例如强心剂、血管扩张剂、抗哮喘剂、抗血栓剂 化合物、平滑肌松弛剂和抗抑郁剂。例如，万艾可，一种 PDE 5抑制剂是一种治疗男性勃起功能障碍处方药 患者该提案旨在表征底物特异性， 抑制剂选择性的结晶学方法。具体目标是 确定PDE及其与抑制剂的复合物的晶体结构 包括（1）PDE 4 B的催化结构域，（2）PDE 4 B的催化结构域 与抑制剂咯利普兰和碘化cAMP复合，（3）全长 PDE 4D及其与抑制剂咯利普兰和登丁茶碱的复合物，以及（4） PDE 3的催化结构域及其与环己酰胺的复合物。的结构 将揭示抑制剂结合在活性 并提供了对催化机理深入了解。将cGMP对接到活性 PDE 4 B的位点，连同PDE 4 B-cAMP类似物的结构， 光对底物的特异性。结构比较 PDE抑制的复合物将揭示抑制的选择性， 不同系列的PDE，从而为设计提供了结构基础， 选择性药物结构将由多个同晶 置换、多波长异常衍射或分子置换。 结构模型将用程序O建立，并由 程序CNS。

DESCRIPTION: (Verbatim from the Applicant's Abstract) Cyclic nucleotide phosphodiesterase (PDE) catalyzes the hydrolysis of adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP) to produce, respectively, 5'-AMP and 5'-GMP. PDE is a key enzyme to control cellular concentrations of cAMP that is known as "second messenger" and mediates the response of cells to a wide variety of hormones and neurotransmitters. Ten families and twenty two subtypes of human PDE have been identified. The mRNAs of the 22 subtype PDEs are further spliced to generate over 60 isoforms of PDE. The distinct isoforms of PDE are located in different cellular compartments and possess different specificity of substrate. These two features of PDE have attracted great attention from pharmaceutical companies in the past decade. Many selective PDE inhibitors have been studied as therapeutic agents such as cardiotonic agents, vasodilators, antiasthma, atithrombic compounds, smooth muscle relaxants and antidepressants. For example, VIAGRA, an inhibitor of PDE5, is a prescription drug for erectile dysfunction of male patients. This proposal aims at characterization of substrate specificity and inhibitor selectivity by the approach of crystallography. The specific aims are to determine crystal structures of PDEs and their complexes with inhibitors including (1) the catalytic domain of PDE4B, (2) the catalytic domain of PDE4B complexed with the inhibitors rolipram and iodonated cAMP, (3) full length PDE4D and its complexes with the inhibitors rolipram and denbufylline and (4) the catalytic domain of PDE3 and its complex with cilostamide. The structures in this proposal will reveal the details of inhibitor binding at the active site and provide insight into catalytic mechanism. Docking cGMP into the active site of PDE4B, together with the structure of PDE4B-cAMP analog, will shed light on the substrate specificity. Comparison of the structures of PDE-inhibited complexes will shed light on the selectivity of inhibition of different families of PDE, and thus provide a structural basis for design of selective drugs. The structures will be determined by multiple isomorphous replacement, multiwavelength anomalous diffraction, or molecular replacement. The structural models will be built with the program O and refined by the program CNS.