Hydropathic analysis and mutagenesis of the catalytic domain of the cGMP-binding cGMP-specific phosphodiesterase (PDE5). cGMP versus cAMP substrate selectivity.
Hydropathic analysis and mutagenesis of the catalytic domain of the cGMP-binding cGMP-specific phosphodiesterase (PDE5). cGMP versus cAMP substrate selectivity.
复制标题
cGMP 结合 cGMP 特异性磷酸二酯酶 (PDE5) 催化结构域的水路分析和诱变。
DOI：
10.1021/bi972448r
复制
发表时间：
1998
期刊：
影响因子：
2.9
通讯作者：
Corbin,JD
中科院分区：
文献类型：
--
作者：
Turko,IV;Francis,SH;Corbin,JD
研究了cGMP结合cGMP特异性磷酸二酯酶(BTPDE5A1或PDE5)催化部位cGMP和cAMP的区别机制。对不同家族PDEs催化结构域的疏水性分析表明，PDEs的底物选择性可能是由于保守的Glu周围短片段中的疏水/亲水残基模式所致，这被证明是PDE5催化域中cGMP结合的关键。这意味着PDE5的底物选择性可以通过将该片段中cGMP特异的PDE中保守的残基替换为cAMP特异的PDE中相应位置的保守残基来改变。A769T/L771R、W762L/Q765Y和W762L/Q765Y/A769T/L771R突变体PDE5在High Five细胞中表达，并与野生型PDE5的底物选择性进行比较。结果表明，PDE5的底物结合部位既含有与cGMP结合的正性元件，也含有阻碍cAMP结合的负性元件，将PDE5的4个残基替换为相应位置的残基，可使PDE5的cGMP/cAMP选择性提高106倍。
The mechanism of discrimination between cGMP and cAMP in the catalytic site of the cGMP-binding cGMP-specific phosphodiesterase (BTPDE5A1 or PDE5) has been investigated. A hydropathy analysis of the catalytic domains of different families of PDEs suggests that substrate selectivity of PDEs could result from the pattern of hydrophobic/hydrophilic residues in a short segment surrounding a conserved Glu that has been shown to be critical for cGMP binding in the catalytic domain of PDE5. This implies that the substrate selectivity of PDE5 could be altered by replacing the residues within this segment that are conserved in cGMP-specific PDEs with the conserved residues in the corresponding positions of cAMP-specific PDEs. The A769T/L771R, W762L/Q765Y, and W762L/Q765Y/A769T/L771R mutant PDE5s were expressed in High Five cells, and their substrate selectivities were compared with that of wild-type PDE5. The results indicate that the substrate-binding site of PDE5 contains positive elements for accommodating cGMP, as well as negative elements that discriminate against binding of cAMP, and that the cGMP/cAMP selectivity of PDE5 can be shifted 106-fold by substituting four residues of PDE5 with the residues in the corresponding positions of PDE4.