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首页> 外文期刊>Molecular and Cellular Biology >Cloning and expression of cDNA for a human low-Km, rolipram-sensitive cyclic AMP phosphodiesterase.
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Cloning and expression of cDNA for a human low-Km, rolipram-sensitive cyclic AMP phosphodiesterase.

机译：人低Km，咯利普兰敏感的环状AMP磷酸二酯酶的cDNA的克隆和表达。

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We have isolated cDNA clones representing cyclic AMP (cAMP)-specific phosphodiesterases (PDEases) from a human monocyte cDNA library. One cDNA clone (hPDE-1) defines a large open reading frame of ca. 2.1 kilobases, predicting a 686-amino-acid, ca. 77-kilodalton protein which contains significant homology to both rat brain and Drosophila cAMP PDEases, especially within an internal conserved domain of ca. 270 residues. Amino acid sequence divergence exists at the NH2 terminus and also within a 40- to 100-residue domain near the COOH-terminal end. hPDE-1 hybridizes to a major 4.8-kilobase mRNA transcript from both human monocytes and placenta. The coding region of hPDE-1 was engineered for expression in COS-1 cells, resulting in the overproduction of cAMP PDEase activity. The hPDE-1 recombinant gene product was identified as a low-Km cAMP phosphodiesterase on the basis of several biochemical properties including selective inhibition by the antidepressant drug rolipram. Known inhibitors of other PDEases (cGMP-specific PDEase, cGMP-inhibited PDEase) had little or no effect on the hPDE-1 recombinant gene product. Human genomic Southern blot analysis suggests that this enzyme is likely to be encoded by a single gene. The presence of the enzyme in monocytes may be important for cell function in inflammation. Rolipram sensitivity, coupled with homology to the Drosophila cAMP PDEase, which is required for learning and memory in flies, suggests an additional function for this enzyme in neurobiochemistry.
机译：我们已经从人单核细胞cDNA文库中分离出代表环AMP（cAMP）特异性磷酸二酯酶（PDEase）的cDNA克隆。一个cDNA克隆（hPDE-1）定义了ca的大型开放阅读框。 2.1千碱基，预测约686个氨基酸。 77-千达尔顿蛋白，与大鼠脑和果蝇cAMP PDEase都具有显着同源性，尤其是在ca的内部保守域内。 270个残基。氨基酸序列差异存在于NH2末端，也位于COOH末端附近的40至100残基域内。 hPDE-1与人单核细胞和胎盘中的主要4.8-kilobase mRNA转录物杂交。 hPDE-1的编码区经过改造，可在COS-1细胞中表达，从而导致cAMP PDEase活性过度产生。基于几种生化特性，包括抗抑郁药咯利普兰的选择性抑制，hPDE-1重组基因产物被鉴定为低Km cAMP磷酸二酯酶。其他PDEase的已知抑制剂（cGMP特异性PDEase，cGMP抑制的PDEase）对hPDE-1重组基因产物几乎没有影响。人基因组Southern印迹分析表明该酶可能由单个基因编码。单核细胞中酶的存在对于炎症中的细胞功能可能很重要。 Rolipram的敏感性以及果蝇cAMP PDEase的同源性（果蝇学习和记忆所必需）表明该酶在神经生物化学中具有其他功能。

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