Characterization of PPIB interaction in the P3H1 ternary complex and implications for its pathological mutations
Characterization of PPIB interaction in the P3H1 ternary complex and implications for its pathological mutations
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P3H1 三元复合物中 PPIB 相互作用的表征及其病理突变的影响
DOI：
10.1007/s00018-019-03102-8
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发表时间：
2019-10-01
影响因子：
8
通讯作者：
Zhou, Aiwu
中科院分区：
文献类型：
--
作者：
Wu, Jiawei;Zhang, Wenting;Zhou, Aiwu
P3 H1/CRTAP/PPIB复合物对于内质网（ER）中脯氨酰3-羟基化和前胶原折叠是必需的。这种三元复合物的任何组分的缺乏都与胶原蛋白的错误折叠和骨形成的开始有关。然而，很少的结构信息是如何组装和保留在ER的三元复合物。在这里，我们评估了P3 H1的KDEL序列的作用，并探讨了PPIB在复合物中的空间相互作用。我们表明，KDEL序列是必不可少的保留P3 H1复杂的ER。其去除导致P3 H1和CRTAP共分泌出细胞，这是通过P3 H1 N-末端结构域与CRTAP结合介导的。分泌的P3 H1/CRTAP可以很容易地结合PPIB，其C-末端靠近三元复合物中的PPIB。半胱氨酸修饰，交联，和质谱实验确定PPIB表面残基参与的复杂的形成，并表明，PPIB的表面被广泛覆盖的P3 H1和CRTAP的结合。最重要的是，我们证明了结合界面上的一个疾病相关的病理性PPIB突变不影响PPIB脯氨酰异构酶活性，但破坏了P3 H1/CRTAP/PPIB三元复合物的形成。这表明P3 H1三元复合物的完整性缺陷与病理性胶原错误折叠有关。总之，这些结果提供了新的结构信息，PPIB如何与P3 H1复合物的其他成分相互作用，并表明P3 H1复合物的完整性是必要的适当的胶原蛋白形成。
The P3H1/CRTAP/PPIB complex is essential for prolyl 3-hydroxylation and folding of procollagens in the endoplasmic reticulum (ER). Deficiency in any component of this ternary complex is associated with the misfolding of collagen and the onset of osteogenesis imperfecta. However, little structure information is available about how this ternary complex is assembled and retained in the ER. Here, we assessed the role of the KDEL sequence of P3H1 and probed the spatial interactions of PPIB in the complex. We show that the KDEL sequence is essential for retaining the P3H1 complex in the ER. Its removal resulted in co-secretion of P3H1 and CRTAP out of the cell, which was mediated by the binding of P3H1 N-terminal domain with CRTAP. The secreted P3H1/CRTAP can readily bind PPIB with their C-termini close to PPIB in the ternary complex. Cysteine modification, crosslinking, and mass spectrometry experiments identified PPIB surface residues involved in the complex formation, and showed that the surface of PPIB is extensively covered by the binding of P3H1 and CRTAP. Most importantly, we demonstrated that one disease-associated pathological PPIB mutation on the binding interface did not affect the PPIB prolyl-isomerase activity, but disrupted the formation of P3H1/CRTAP/PPIB ternary complex. This suggests that defects in the integrity of the P3H1 ternary complex are associated with pathological collagen misfolding. Taken together, these results provide novel structural information on how PPIB interacts with other components of the P3H1 complex and indicate that the integrity of P3H1 complex is required for proper collagen formation.