ISOLATION AND USES OF CGMP PHOSPHODIESTERASE CDNAS

CGMP 磷酸二酯酶 CDNAS 的分离和用途

• 批准号: 3306537
• 负责人: RICHARD Harry KESSIN
• 金额: $ 26.36万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-02-01 至 1996-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3306537
• 关键词: Dictyostelium; biological signal transduction; cell motility; complementary DNA; cyclic AMP; cyclic GMP; endonuclease; enzyme induction /repression; enzyme mechanism; gene expression; genetic manipulation; guanine nucleotide binding protein; histogenesis; isozymes; molecular cloning; nucleic acid sequence; phosphodiesterases; polymerase chain reaction; protein purification; pulsed field gel electrophoresis; radioimmunoassay

The cyclic nucleotide phosphodiesterases are crucial components in the regulation of the signal transduction cascades mediated by cAMP and cGMP. The size of this family of enzymes and the complexity of their regulation may not be adequately appreciated. The importance of studying them stems from their intrinsic biochemical and biological interest and from the fact that they are the targets of anti-asthmatic, anti-thrombolytic, cardiotonic, and anti-depressant drugs. Although they have been studied at the biochemical level for many years, the cDNAs and genes of some isoforms have been difficult to clone and this has now become a necessity. We have developed a genetic selection procedure that allows recovery of cDNAs coding for these enzymes. Conditions have been created in which lambda lysogens of E. coli that are capable of expressing a functional cyclic nucleotide or cGMP phosphodiesterase can grow. This method has been used to recover cDNAs that may code for a cGMP stimulated cGMP specific phosphodiesterase from Dictyostelium discoideum. The cGMP mediated signal transduction cascade and this phosphodiesterase regulate the motility of this organism and will be used to ask questions about how cGMP mediated signal transduction controls mobilization of the cytoskeleton and other developmentally important events. The same selection procedure will be improved and extended to recover selected mammalian phosphodiesterase cDNAs and to study their molecular, biochemical, and pharmacological properties.

环核苷酸磷酸二酯酶是环核苷酸磷酸二酯酶中的关键组分。 调节cAMP和cGMP介导的信号转导级联。 这个酶家族的规模和它们调控的复杂性 可能没有得到充分的重视。 研究它们的重要性源于 从他们内在的生物化学和生物学兴趣， 它们是抗哮喘、抗血栓溶解 强心剂和抗心律失常药。 尽管它们已经在 多年来的生化水平，一些异构体的cDNA和基因， 很难克隆，现在这已经成为一种必要。 我们开发了一种基因选择程序， 编码这些酶的cDNA。 已经创造了条件， λ溶原菌。能够表达功能性 环核苷酸或cGMP磷酸二酯酶可以生长。 该方法已 用于回收可能编码cGMP刺激的cGMP特异性的cDNA， 来自盘基网柄藻的磷酸二酯酶。 cGMP介导信号 转导级联和这种磷酸二酯酶调节运动的 这种生物体，并将用于询问有关cGMP如何介导的问题 信号转导控制细胞骨架和其他细胞的移动， 重要的发展事件。 同样的选择程序将 改进和扩展以回收选定的哺乳动物磷酸二酯酶cDNA 并研究其分子、生物化学和药理学特性。