Reversibly bound chloride in the atrial natriuretic peptide receptor hormone-binding domain: possible allosteric regulation and a conserved structural motif for the chloride-binding site.
Reversibly bound chloride in the atrial natriuretic peptide receptor hormone-binding domain: possible allosteric regulation and a conserved structural motif for the chloride-binding site.
复制标题
心房钠尿肽受体激素结合域中可逆结合的氯：可能的变构调节和氯结合位点的保守结构基序。
DOI：
10.1002/pro.332
复制
发表时间：
2010
期刊：
影响因子：
--
通讯作者：
Misono,KunioS
中科院分区：
文献类型：
--
作者：
Ogawa,Haruo;Qiu,Yue;Philo,JohnS;Arakawa,Tsutomu;Ogata,CraigM;Misono,KunioS
心房钠尿肽（ANP）与其受体的结合需要氯化物，并且它是氯化物浓度依赖性的。 ANP 受体 (ANPR) 的胞外结构域 (ECD) 在 ANP 结合位点附近含有一个氯，表明可能具有调节作用。然而，结合的氯完全埋藏在多肽折叠中，其功能作用仍不清楚。在这里，我们已经证实氯对于 ANP 与重组 ECD 或 CHO 细胞中表达的全长 ANPR 结合是必需的。不含氯化物的 ECD (ECD(−)) 不结合 ANP。通过添加溴化物或氯化物，其结合活性完全恢复。溴化物结合 ECD 的新 X 射线结构与氯离子结合 ECD 基本相同。此外，在apo和ANP结合结构中，溴原子位于与氯原子相同的位置，表明卤化物结合是可交换和可逆的。远紫外 CD 和热展开数据表明 ECD(−) 在很大程度上保留了天然结构。在没有氯化物的情况下的沉降平衡表明，ECD(−) 形成紧密相关的二聚体，可能阻止 ANP 结合所需的两个单体的结构重排。 ANPR 中氯离子结合位点的一级和三级结构在受体鸟苷酸环化酶和代谢型谷氨酸受体中高度保守。氯依赖性 ANP 结合、可逆氯结合和高度保守的氯结合位点基序表明受体结合氯具有调节作用。 ANPR 的氯化物依赖性调节可能在肾脏中发挥作用，调节 ANP 诱导的尿钠排泄。
The binding of atrial natriuretic peptide (ANP) to its receptor requires chloride, and it is chloride concentration dependent. The extracellular domain (ECD) of the ANP receptor (ANPR) contains a chloride near the ANP‐binding site, suggesting a possible regulatory role. The bound chloride, however, is completely buried in the polypeptide fold, and its functional role has remained unclear. Here, we have confirmed that chloride is necessary for ANP binding to the recombinant ECD or the full‐length ANPR expressed in CHO cells. ECD without chloride (ECD(−)) did not bind ANP. Its binding activity was fully restored by bromide or chloride addition. A new X‐ray structure of the bromide‐bound ECD is essentially identical to that of the chloride‐bound ECD. Furthermore, bromide atoms are localized at the same positions as chloride atoms both in the apo and in the ANP‐bound structures, indicating exchangeable and reversible halide binding. Far‐UV CD and thermal unfolding data show that ECD(−) largely retains the native structure. Sedimentation equilibrium in the absence of chloride shows that ECD(−) forms a strongly associated dimer, possibly preventing the structural rearrangement of the two monomers that is necessary for ANP binding. The primary and tertiary structures of the chloride‐binding site in ANPR are highly conserved among receptor‐guanylate cyclases and metabotropic glutamate receptors. The chloride‐dependent ANP binding, reversible chloride binding, and the highly conserved chloride‐binding site motif suggest a regulatory role for the receptor bound chloride. Chloride‐dependent regulation of ANPR may operate in the kidney, modulating ANP‐induced natriuresis.