Cryo‐EM analysis provides new mechanistic insight into ATP binding to Ca2+‐ATPase SERCA2b
Cryo‐EM analysis provides new mechanistic insight into ATP binding to Ca2+‐ATPase SERCA2b
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DOI：
10.15252/embj.2021108482
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发表时间：
2021-08
期刊：
影响因子：
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通讯作者：
Yuxia Zhang;Satoshi Watanabe;A. Tsutsumi;H. Kadokura;M. Kikkawa;K. Inaba
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作者：
Yuxia Zhang;Satoshi Watanabe;A. Tsutsumi;H. Kadokura;M. Kikkawa;K. Inaba
肌浆/内质网 Ca2+-ATP 酶 (SERCA) 2b 是普遍存在的 SERCA 家族成员，负责将 Ca2+ 从细胞质摄取到内质网。在此，我们展示了处于 E1·2Ca2+ 状态的人 SERCA2b 的 3.3 Å 分辨率冷冻电子显微镜 (cryo-EM) 结构，揭示了 Ca2+ 结合 SERCA2b 的新构象，其胞质结构域的排列比之前报道的 Ca2+ 结合 SERCA1a 的晶体结构更接近。冷冻电镜图 3D 分类生成的多种构象反映了这种状态下胞质域的内在动态性质。值得注意的是，E1·2Ca2+状态下SERCA2b的ATP结合残基与E1·2Ca2+-ATP状态下的ATP结合残基位于相似的位置。因此，冷冻电镜结构可能代表了 ATP 结合之前的预形成状态。一致地，具有锁定闭合胞质结构域排列的域间二硫桥的 SERCA2b 突变体在 Ca2+ 存在下表现出显着的自磷酸化活性。我们提出了一种 ATP 与 SERCA2b 结合的新机制。
Sarco/endoplasmic reticulum Ca2+‐ATPase (SERCA) 2b is a ubiquitous SERCA family member that conducts Ca2+ uptake from the cytosol to the ER. Herein, we present a 3.3 Å resolution cryo‐electron microscopy (cryo‐EM) structure of human SERCA2b in the E1·2Ca2+ state, revealing a new conformation for Ca2+‐bound SERCA2b with a much closer arrangement of cytosolic domains than in the previously reported crystal structure of Ca2+‐bound SERCA1a. Multiple conformations generated by 3D classification of cryo‐EM maps reflect the intrinsically dynamic nature of the cytosolic domains in this state. Notably, ATP binding residues of SERCA2b in the E1·2Ca2+ state are located at similar positions to those in the E1·2Ca2+‐ATP state; hence, the cryo‐EM structure likely represents a preformed state immediately prior to ATP binding. Consistently, a SERCA2b mutant with an interdomain disulfide bridge that locks the closed cytosolic domain arrangement displayed significant autophosphorylation activity in the presence of Ca2+. We propose a novel mechanism of ATP binding to SERCA2b.