GTP BINDING PROTEINS AND ADENYLATE CYCLASE

GTP 结合蛋白和腺苷酸环化酶

• 批准号: 4694493
• 负责人: S C TSAI
• 金额: --
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/4694493
• 关键词: adenosine diphosphate; adenylate cyclase; bacterial toxins; centrifugation; chromatography; enzyme complex; enzyme reconstitution; fluorine; gel electrophoresis; guanosine triphosphate; hormone regulation /control mechanism; pertussis; phosphodiesterases; radiotracer; rhodopsin; ribose; vanadium

Adenylate cyclase systems consist of inhibitory and stimulatory receptors linked to a catalytic unit through inhibitory and stimulatory proteins, Gi and Gs, respectively. Similarly, the retinal photon receptor rhodopsin is linked with a phosphodiesterase through transducin (T), which is similar to Gs and Gi. Each has a guanyl nucleotide-binding Alpha subunit (45 kDa Gs, 41 kDa Gi, and 39 kDa T) and Beta, Gamma subunit (35 and l0 kDa). Gs and Gi can be activated by NaF with AlCl3 and by decavadanate, as well as by agonists acting through specific receptors. The effects of NaF with AlCl3 and decavanadate on transducin were investigated in a reconstituted system consisting of the purified subunits of transducin (TAlpha, TBeta Gamma) and rhodopsin in prhosphatidylcholine vesicles. NaF noncompetitively inhibited the GTPase activity of TAlpha in a concentration- and time-dependent manner. Inhibition by NaF was enhanced synergistically by AlCl3 which alone only slighly inhibited GTPase activity. None of the other anions tested reproduced the effect of fluoride. Fluoride inhibited [3H]GppNHp binding to TAlpha and release of bound GDP. Vanadate (decameric) inhibited [3H]GTP binding to TAlpha and GTP hydrolysis in a concentration-dependent manner with maximal inhibition of approximately 90% at 3-5 mM. Vanadate also inhibited release of bound GDP. The ADP-ribosylation of TAlpha by pertussis toxin and binding of TAlpha to rhodopsin, both of which are enhanced in the presence of TBetaGamma, were inhibited by NaF with AlCl3 and by vanadate. These findings are consistent with the conclusion that NaF plus AlCl3 or vanadate can cause the dissociation of TAlpha from TBetaGamma, resulting in the inhibition of GDP-GTP exchange and thereby GTP hydrolysis. Adenylate cyclase activation and inhibition presumably result from similar effects of NaF plus AlCl3 and vanadate on Gs and Gi.

腺苷酸环化酶系统由抑制性和刺激性受体组成 通过抑制性和刺激性蛋白质Gi与催化单位连接 G，分别。 类似地，视网膜光子受体视紫红质是 通过转导素（T）与磷酸二酯酶连接，类似于 G和Gi。 每个都有一个鸟苷酸结合α亚基（45 kDa Gs， 41 kDa Gi和39 kDa T）和β、γ亚基（35和10 kDa）。 Gs和 Gi可以被NaF和AlCl 3激活，也可以被十钒酸盐激活， 通过特定受体起作用的激动剂。 NaF与AlCl 3的作用 和十钒酸钠对转导蛋白的影响 由纯化的转导素亚基（T α，T β γ）组成， 在磷脂酰胆碱囊泡中的视紫红质。 NaF非竞争性抑制 TAlpha浓度和时间依赖性GT3活性 方式 NaF的抑制作用被AlCl 3协同增强， 单独仅轻微抑制GT3活性。 其他的阴离子 试验再现了氟化物的作用。氟抑制[3 H]GppNHp 与TAlpha结合并释放结合的GDP。 钒酸盐（十聚体）抑制 [3 H]GTP与TAlpha的结合和GTP水解呈浓度依赖性 在3-5 mM时，最大抑制率约为90%。 也抑制了绑定GDP的释放。 TAlpha的ADP-核糖基化 百日咳毒素和TAlpha与视紫红质的结合，两者都是 在TBetaGamma存在下增强，被NaF和AlCl 3抑制 和钒酸盐。 这些发现与以下结论一致： NaF加上AlCl 3或钒酸盐可以导致TAlpha从 TBetaGamma，导致GDP-GTP交换的抑制，从而抑制GTP 水解 腺苷酸环化酶的激活和抑制可能导致 NaF加AlCl 3和钒酸盐对Gs和Gi的类似作用。