GTP BINDING PROTEINS AND ADENYLATE CYCLASE

GTP 结合蛋白和腺苷酸环化酶

• 批准号: 3919997
• 负责人: M NODA
• 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/3919997
• 关键词: ADP ribosylation; G protein; adenylate cyclase; binding proteins; brain; cholera toxin; cow; enzyme complex; enzyme induction /repression; enzyme mechanism; enzyme structure; enzyme substrate; guanidines; guanosine triphosphate; human tissue; nicotinamide adenine dinucleotide; peptides; pertussis toxin; ribose

Choleragen (cholera toxin), the agent responsible for the devastating diarrheal disease characteristic of cholera, causes its effects on cells by activating the hormone-sensitive adenylyl cyclase. Activation results from the ADP-ribosylation of Gsa, a guanine nucleotide-binding regulatory protein of the cyclase complex. The toxin can also ADP-ribosylate other proteins as well as simple guanidino compounds such as agmatine. ADP-ribosylation of Gs alpha and other substrates is enhanced by so-called ADP- ribosylation factors (ARFs), a family of approximately 19 kDa soluble and membrane guanine nucleotide-binding proteins that have been purified from bovine brain. In the presence of GTP, but not GDP, ARF interacts with the toxin resulting in a decrease in apparent Kms for both NAD and ADP-ribose acceptor with no significant effect on Vmax. Activation of toxin by ARF was increased approximately 4-fold by 0.003% sodium dodecyl sulfate (SDS), which had little or no effect on activity in the absence of ARF; 0.01% SDS abolished toxin activity with or without ARF. In the presence of SDS at stimulatory concentrations, ARF further increased affinities for NAD and agmatine and, in addition, increased Vmax. Stimulatory effects of ARF and SDS were observed in the presence of dithiothreitol with toxin A subunit and in the absence of dithiothreitol, with the reduced and alkylated toxin A1 peptide. Thus, it appears that ARF and SDS do not activate by facilitating release of the toxin catalytic unit. Rather, ARF interacts directly with the catalytic A1 peptide and may thus potentiate the pathogenicity of cholera toxin.

霍乱原（霍乱毒素），负责 一种以霍乱为特征的毁灭性霍乱， 通过激活对腺苷酸敏感的腺苷酸 环化酶。 激活的结果从ADP核糖基化的Gsa， 环化酶鸟嘌呤核苷酸结合调节蛋白 复杂. 该毒素也可以ADP核糖基化其他蛋白质 作为简单的胍基化合物如胍丁胺。 ADP-核糖基化 Gs α和其他底物的作用被所谓的ADP增强， 核糖基化因子（ARF），一个约19 kDa的家族， 可溶性和膜鸟嘌呤核苷酸结合蛋白， 是从牛脑中提纯出来的 在GTP的存在下，但不是 GDP、ARF与毒素相互作用，导致 NAD和ADP-核糖受体的表观Km，无 对Vmax有显著影响。 ARF对毒素的激活作用 添加0.003%十二烷基硫酸钠可使其增加约4倍 (SDS)，在没有 ARF; 0.01%SDS在有或无ARF的情况下均能消除毒素活性。 在 在刺激浓度SDS存在下，ARF进一步 对NAD和胍丁胺的亲和力增加，此外， 增加Vmax。 观察ARF和SDS的刺激效应 在具有毒素A亚基的二硫苏糖醇的存在下， 不含二硫苏糖醇，含还原和烷基化毒素A1 肽。 因此，似乎ARF和SDS不被激活， 促进毒素催化单元的释放。 相反， 直接与催化A1肽相互作用， 增强霍乱毒素的致病性。