Purification and properties of cyclic phosphodiesterase: 3'-nucleotidase, a periplasmic enzyme of Haemophilus influenzae.
Purification and properties of cyclic phosphodiesterase: 3'-nucleotidase, a periplasmic enzyme of Haemophilus influenzae.
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环磷酸二酯酶的纯化和性质：3-核苷酸酶，流感嗜血杆菌的周质酶。
DOI：
10.1016/0003-9861(72)90406-7
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发表时间：
1972
影响因子：
3.9
通讯作者：
J. J. Scocca
中科院分区：
文献类型：
--
作者：
J. Rodden;J. J. Scocca
环状磷酸二酯酶：流感嗜血杆菌的3′-核苷酸酶催化2′，3 ′-环核苷酸裂解成3′-单酯，并水解成核苷和无机磷酸盐。它还水解人工底物双（对硝基苯基）磷酸酯，产生对硝基苯酚和对硝基苯基磷酸酯。该酶位于H.流感嗜血杆菌中的这种酶，可以通过使细胞经受渗透压休克或通过将细胞转化为原生质球而选择性地释放。通过在Tris、EDTA和Nonidet P-40的溶液中孵育细胞来影响流感病毒;将酶进一步纯化至表观电泳均一性。该酶来自H.在没有任何金属离子的情况下，流感病毒水解3′-核苷酸，尽管Co ~（2+）和Ca ~（2+）刺激双（对-硝基苯）磷酸的裂解，我们观察到2′-、3′-和5′-腺苷单磷酸对双（对-硝基苯）磷酸的裂解有明显的竞争性抑制。2′-和5′-核苷酸也抑制环磷酸二酯酶和3′-核苷酸酶的活性。这些结果表明，核苷酸底物和人工显色底物与相同的活性位点相互作用。
The cyclic phosphodiesterase: 3′-nucleotidase ofHaemophilus influenzaecatalyzes the cleavage of 2′,3′-cyclic nucleotides to 3′-monoesters, and the hydrolysis of the resulting 3′-nucleotides to nucleosides and inorganic phosphate. It also hydrolyzes the artificial substrate bis(p-nitrophenyl)phosphate, yieldingp-nitrophenol andp-nitrophenyl phosphate. The enzyme is located in the periplasmic space ofH. influenzae, and can be selectively released by subjecting the cells to osmotic shock, or by converting the cells to spheroplasts.The selective removal of the enzyme fromH. influenzaewas effected by incubating the cells in a solution of Tris, EDTA, and Nonidet P-40; the enzyme was further purified to apparent electrophoretic homogeneity. The enzyme fromH. influenzaehydrolyses 3′-nucleotides in the absence of any added metal ion, although Co2+and Ca2+stimulate the cleavage of bis(p-nitropheny])phosphate.We have observed a marked competitive inhibition of bis(p-nitrophenyl)phosphate cleavage by 2′-, 3′-, and 5′-adenosine monophosphate. The 2′- and 5′-nucleotides also inhibit the cyclic phosphodiesterase and 3′-nucleotidase activities. These results indicate that nucleotide substrates and the artificial chromogenic substrate interact with the same active site.