Ecto-nucleotide pyrophosphatase/phosphodiesterase as part of a multiple system for nucleotide hydrolysis by platelets from rats:: Kinetic characterization and biochemical properties
Ecto-nucleotide pyrophosphatase/phosphodiesterase as part of a multiple system for nucleotide hydrolysis by platelets from rats:: Kinetic characterization and biochemical properties
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DOI：
10.1080/09537100500246641
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发表时间：
2006-03-01
期刊：
影响因子：
3.3
通讯作者：
Sarkis, JJF
中科院分区：
文献类型：
--
作者：
Fürstenau, CR;Trentin, DD;Sarkis, JJF
在这项研究中，我们描述了大鼠血小板中的外核苷酸焦磷酸酶/磷酸二酯酶（E-NPP）活性。利用对硝基苯基5′-胸苷单磷酸（p-Nph-5′-TMP）作为E-NPP的底物，我们证明了一种酶的活性具有E-NPP的主要生化特性：碱性pH依赖性、二价阳离子依赖性和金属离子螯合剂对活性的阻断。p-Nph-5′- tmp水解的K-m和V-max值分别为106 +/- 18 μ M和3.44 +/- 0.18 nmol对硝基苯酚/min/mg（平均+/- SD， n = 5）。我们假设E-NPP与外核苷三磷酸二磷酸水解酶和外5'-核苷酸酶在血小板表面共定位，作为核苷酸水解的多系统的一部分，因为它们可以在不同的生理条件下起作用，并且可以不同地调节。因此，0.25 mM苏拉明抑制p-Nph-5′-TMP、ATP和ADP的水解，而0.5 mM AMP仅抑制p-Nph-5′-TMP的水解。此外，5.0、10和20 mM叠氮化钠仅能抑制ATP和ADP的水解。血管紧张素II （5.0 nM和10 nM）仅影响ADP水解。氯化钆（0.2和0.5 mM）对ATP和ADP的水解有较强的抑制作用。这里描述的E-NPP代表了对血小板嘌呤能信号控制的新见解。
In this study, we describe an ecto-nucleotide pyrophosphatase/phosphodiesterase (E-NPP) activity in rat platelets. Using p-nitrophenyl 5'-thymidine monophosphate (p-Nph-5'-TMP) as a substrate for E-NPP, we demonstrate an enzyme activity that shares the major biochemical properties described for E-NPPs: alkaline pH dependence, divalent cation dependence and blockade of activity by metal ion chelator. K-m and V-max values for p-Nph-5'-TMP hydrolysis were found to be 106 +/- 18 mu M and 3.44 +/- 0.18 nmol p-nitrophenol/min/mg ( mean +/- SD, n = 5). We hypothesize that an E-NPP is co-localized with an ecto-nucleoside triphosphate diphosphohydrolase and an ecto-5'-nucleotidase on the platelet surface, as part of a multiple system for nucleotide hydrolysis, since they can act under distinct physiological conditions and can be differently regulated. Thus, 0.25 mM suramin inhibited p-Nph-5'-TMP, ATP and ADP hydrolysis, while 0.5 mM AMP decreased only p-Nph-5'-TMP hydrolysis. Besides, 5.0, 10 and 20 mM sodium azide just inhibited ATP and ADP hydrolysis. Angiotensin II (5.0 and 10 nM) affected only ADP hydrolysis. Gadolinium chloride (0.2 and 0.5 mM) strongly inhibited the ATP and ADP hydrolysis. The E-NPP described here represents a novel insight into the control of platelet purinergic signaling.