Structural insights for activation of retinal guanylate cyclase by GCAP1.
Structural insights for activation of retinal guanylate cyclase by GCAP1.
复制标题
DOI：
10.1371/journal.pone.0081822
复制
发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Ames JB
中科院分区：
文献类型：
--
作者：
Lim S;Peshenko IV;Dizhoor AM;Ames JB
观酰基环化酶激活蛋白1 （GCAP1）是钙调蛋白超家族中神经元钙传感器（NCS）亚类的成员，在光感受器细胞的光激活下，赋予视网膜观酰基环化酶1 (RetGC1) Ca2+敏感激活。在这里，我们提出了核磁共振分配和功能分析，以探测Ca2+依赖的GCAP1结构变化，控制RetGC的激活。获得了GCAP1 Ca2+饱和抑制状态与GCAP1突变体（D144N/D148G，称为EF4mut）的核磁共振分配，后者在EF-hand 4中缺乏Ca2+结合，并模拟了GCAP1的Ca2+游离/Mg2+结合激活状态。Ca2+饱和野生型GCAP1的主链共振核磁共振化学位移总体上与EF4mut相似，表明在Ca2+游离激活剂和Ca2+结合抑制剂状态下，指定残基的主链结构相似。这与Ca2+诱导的化学位移差异形成鲜明对比，因此其他NCS蛋白（包括recoverin和NCS-1）的结构发生了巨大变化。GCAP1与EF4mut化学位移差异最大的是EF4中的残基（S141、K142、V145、N146、G147、G149、E150、L153、E154、M157、E158、Q161、L166），而EF4残基（F140A、K142D、L153R、L166R）的诱变对RetGC1的激活影响不大。EF-hand 1中少数GCAP1残基（K23、T27、G32）也表现出较大的化学位移差异，其中两个突变（K23D和G32N）均降低了RetGC的激活，这与EF1的功能构象变化一致。GCAP1结构域界面上的残基（V77, A78, L82）具有交换展宽的核磁共振，表明这些残基可能具有构象动力学，这与先前的研究结果一致，表明这些残基位于激活RetGC1所必需的区域。
Guanylyl cyclase activating protein 1 (GCAP1), a member of the neuronal calcium sensor (NCS) subclass of the calmodulin superfamily, confers Ca2+-sensitive activation of retinal guanylyl cyclase 1 (RetGC1) upon light activation of photoreceptor cells. Here we present NMR assignments and functional analysis to probe Ca2+-dependent structural changes in GCAP1 that control activation of RetGC. NMR assignments were obtained for both the Ca2+-saturated inhibitory state of GCAP1 versus a GCAP1 mutant (D144N/D148G, called EF4mut), which lacks Ca2+ binding in EF-hand 4 and models the Ca2+-free/Mg2+-bound activator state of GCAP1. NMR chemical shifts of backbone resonances for Ca2+-saturated wild type GCAP1 are overall similar to those of EF4mut, suggesting a similar main chain structure for assigned residues in both the Ca2+-free activator and Ca2+-bound inhibitor states. This contrasts with large Ca2+-induced chemical shift differences and hence dramatic structural changes seen for other NCS proteins including recoverin and NCS-1. The largest chemical shift differences between GCAP1 and EF4mut are seen for residues in EF4 (S141, K142, V145, N146, G147, G149, E150, L153, E154, M157, E158, Q161, L166), but mutagenesis of EF4 residues (F140A, K142D, L153R, L166R) had little effect on RetGC1 activation. A few GCAP1 residues in EF-hand 1 (K23, T27, G32) also show large chemical shift differences, and two of the mutations (K23D and G32N) each decrease the activation of RetGC, consistent with a functional conformational change in EF1. GCAP1 residues at the domain interface (V77, A78, L82) have NMR resonances that are exchange broadened, suggesting these residues may be conformationally dynamic, consistent with previous studies showing these residues are in a region essential for activating RetGC1.
登录
查看更多内容
影响因子：
4.8
作者：
Dizhoor, AM;Hurley, JB
通讯作者：
Hurley, JB
影响因子：
4.3
作者：
Dizhoor, Alexander M.;Olshevskaya, Elena V.;Peshenko, Igor V.
通讯作者：
Peshenko, Igor V.
影响因子：
34.7
作者：
Burgoyne, Robert D.
通讯作者：
Burgoyne, Robert D.
影响因子：
4.8
作者：
Ames, JB;Tanaka, T;Stryer, L
通讯作者：
Stryer, L
DOI：
10.1073/pnas.93.2.560
发表时间：
1996-01-23
影响因子：
11.1
作者：
Baylor, D
通讯作者：
Baylor, D