Adenosine inhibition of catecholamine-stimulated cardiac membrane adenylate cyclase.
Adenosine inhibition of catecholamine-stimulated cardiac membrane adenylate cyclase.
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DOI：
10.1152/ajpheart.1985.248.5.h737
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发表时间：
1985-05
期刊：
影响因子：
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通讯作者：
D. Lamonica;N. Frohloff;J. G. Dobson
中科院分区：
文献类型：
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作者：
D. Lamonica;N. Frohloff;J. G. Dobson
腺苷抑制酶敏感腺苷酸环化酶活性进行了研究，使用离体心肌膜制备从大鼠心脏。当在膜中测定环化酶活性时，使用[α-32 P]ATP作为底物，10（-5）M腺苷抑制异丙肾上腺素刺激的腺苷酸环化酶活性25%，但不抑制酶的基础活性或氟化物（5 mM）活化。腺苷降低异丙肾上腺素敏感性环化酶活性依赖于GTP，但不能被10（-3）M茶碱阻止。腺苷似乎既不与ATP竞争酶的底物转化位点，也不降低酶的5 '-鸟苷酰亚胺二磷酸活化。由于较低浓度的腺苷对酶活性没有影响，因此腺苷酸环化酶测定中可能存在内源性腺苷。为了验证内源性腺苷的影响，然后将腺苷酸环化酶测定法修改为在腺苷脱氨酶存在下使用[α-32 P]dATP作为底物的2 '-脱氧系统。在该测定系统中，腺苷受体激动剂10（-8）M 2-氯腺苷或苯基异丙基腺苷对异丙肾上腺素刺激的腺苷酸环化酶活性产生的15%抑制作用分别被10（-4）M 8-苯基茶碱或异丁基甲基黄嘌呤（IBMX）阻止。而在这些测定条件下，10（-7）M的2 '，5'-双脱氧腺苷，一种P-位点类似物，不影响酶敏感的环化酶活性。在10（-5）M浓度下，IBMX不能阻止该类似物产生的酶敏感性酶降低35%。这些结果表明，纳摩尔浓度的腺苷类似物与甲基黄嘌呤敏感的腺苷受体，介导的膜脂敏感的腺苷酸环化酶活性的注意相互作用。
Adenosine inhibition of hormone-sensitive adenylate cyclase activity was investigated using isolated myocardial membranes prepared from rat hearts. When cyclase activity was determined in membranes, using [alpha-32P]ATP as substrate, 10(-5) M adenosine inhibited isoproterenol-stimulated adenylate cyclase activity by 25% but did not inhibit basal activity or fluoride (5 mM) activation of the enzyme. The adenosine reduction of isoproterenol-sensitive cyclase activity was dependent on GTP but was not prevented by 10(-3) M theophylline. Adenosine neither appeared to compete with ATP for the substrate converting site of the enzyme nor reduced 5'-guanylyl imidodiphosphate activation of the enzyme. Inasmuch as lower concentrations of adenosine had no influence on enzyme activity, endogenous adenosine may be present in the adenylate cyclase assay. To obviate the effects of endogenous adenosine, the adenylate cyclase assay was then modified to a 2'-deoxy system with [alpha-32P]dATP used as the substrate in the presence of adenosine deaminase. With this assay system, the 15% inhibition of isoproterenol-stimulated adenylate cyclase activity produced by the adenosine receptor agonists, 10(-8) M 2-chloroadenosine or phenylisopropyladenosine, was prevented by 10(-4) M 8-phenyltheophylline or isobutylmethylxanthine (IBMX), respectively. While under these assay conditions, 10(-7) M 2',5'-dideoxyadenosine, a P-site analogue, did not influence the hormone-sensitive cyclase activity. The 35% reduction of the hormone-sensitive enzyme produced by this analogue at 10(-5) M was not prevented by IBMX. These results suggest that nanomolar concentrations of adenosine analogues interact with a methylxanthine-sensitive adenosine receptor that mediates the attention of membrane hormone-sensitive adenylate cyclase activity.