Visual arrestin 1 (Arr1) is an essential protein for termination of the light response in photoreceptors. While mammalian Arr1s form dimers and tetramers at physiological concentrations in vitro, oligomerization in other vertebrates has not been studied. Here we examine self-association of Arr1 from two amphibian species, Xenopus laevis (xArr1), and Ambystoma tigrinum (salArr1). Sedimentation velocity analytical ultracentrifugation showed that xArr1 and salArr1 oligomerization is limited to dimers. The KD for dimer formation was 53 μM for xArr1 and 44 μM for salArr1, similar to the 69 μM KD for bovine Arr1 (bArr1) dimers. Mutations of orthologous amino acids important for mammalian Arr1 oligomerization had no impact on xArr1 dimerization. Crystallography showed that the fold of xArr1 closely resembles that of bArr1 and crystal structures in different space groups revealed two potential xArr1 dimer forms: a symmetrical dimer with a C-domain interface (CC dimer), resembling the bArr1 solution dimer, and an asymmetric dimer with an N-domain/C-domain interface. Mutagenesis of residues predicted to interact in either of these two dimer forms yielded modest reduction in dimer affinity, suggesting that the dimer interfaces compete or are not unique. Indeed, small-angle X-ray scattering and protein painting data were consistent with a symmetric anti-parallel solution dimer (AP dimer) distinct from the assemblies observed by crystallography. Finally, a computational model evaluating xArr1 binding to compartment-specific partners and partitioning based on heterogeneity of available cytoplasmic spaces shows that Arr1 distribution in dark adapted photoreceptors is largely explained by the excluded volume effect together with tuning by oligomerization.

中文翻译：

---

两栖动物 arrestin 1 自缔合和视网膜光感受器动态分布的机制。


视觉抑制蛋白 1 （Arr1） 是终止光感受器中光反应的必需蛋白。虽然哺乳动物 Arr1 在体外形成生理浓度的二聚体和四聚体，但尚未研究其他脊椎动物的寡聚化。在这里，我们检查了来自两个两栖动物物种 Xenopus laevis （xArr1） 和 Ambystoma tigrinum （salArr1） 的 Arr1 的自缔合。沉降速度分析超速离心显示 xArr1 和 salArr1 寡聚化仅限于二聚体。xArr1 形成二聚体的 KD 为 53 μM，salArr1 为 44 μM，与牛 Arr1 （bArr1） 二聚体形成的 KD 为 69 μM。对哺乳动物 Arr1 寡聚化很重要的直系同源氨基酸突变对 xArr1 二聚化没有影响。晶体学表明，xArr1 的折叠与 bArr1 的折叠非常相似，不同空间群中的晶体结构揭示了两种潜在的 xArr1 二聚体形式：具有 C 结构域界面 （CC 二聚体） 的对称二聚体，类似于 bArr1 溶液二聚体，以及具有 N 结构域/C 结构域界面的不对称二聚体。预测以这两种二聚体形式中的任何一种相互作用的残基的诱变导致二聚体亲和力适度降低，表明二聚体界面竞争或不唯一。事实上，小角 X 射线散射和蛋白质绘画数据与对称的反平行溶液二聚体 （AP 二聚体） 一致，这与晶体学观察到的组装体不同。最后，评估 xArr1 与区室特异性伴侣结合和基于可用细胞质空间异质性的分区的计算模型表明，Arr1 在暗适应光感受器中的分布在很大程度上是由排除的体积效应以及寡聚化的调节来解释的。