软骨酸性蛋白-1 (CRTAC1) 是一种分泌性糖蛋白，在神经系统的发育、功能和修复中发挥作用。它与缺血性中风、骨关节炎和（长期）新冠病毒结局有关，并且对癌症和膀胱癌具有抑制活性。 CRTAC1 的结构表征因其形成二硫键连接聚集体的倾向而变得复杂。在这里，我们证明 CRTAC1 被钾离子稳定。使用 X 射线晶体学，我们确定了 CRTAC1 的结构为 1.6 Å。这表明该蛋白质由三结构域折叠组成，包括先前未报道的紧凑型β-螺旋桨-TTR组合，其中TTR的延伸环堵塞了β-螺旋桨核心。观察十种结合离子的电子密度：六种钙、三种钾和一种钠。低钾离子浓度会导致色氨酸环境发生变化，并导致位于 β 叶片上和 β 螺旋桨堵塞 TTR 环中的两个埋藏游离半胱氨酸暴露。在没有钾离子的情况下，将两个游离半胱氨酸突变为丝氨酸可防止共价分子间相互作用，但不会聚集。钾离子结合位点位于 β 螺旋桨的叶片之间，这解释了它们对于 CRTAC1 折叠稳定性的重要性。尽管序列不同，三个钾离子结合位点在结构上相似并且具有 CRTAC 蛋白家族的保守特征。这些对 CRTAC1 稳定性和结构的见解为进一步研究 CRTAC1 在健康和疾病中的功能奠定了基础。版权所有 © 2024 作者。由爱思唯尔有限公司出版。保留所有权利。

Cartilage acidic protein-1 (CRTAC1) is a secreted glycoprotein with roles in development, function and repair of the nervous system. It is linked to ischemic stroke, osteoarthritis and (long) COVID outcomes, and has suppressive activity in carcinoma and bladder cancer. Structural characterization of CRTAC1 has been complicated by its tendency to form disulfide-linked aggregates. Here, we show that CRTAC1 is stabilized by potassium ions. Using x-ray crystallography, we determined the structure of CRTAC1 to 1.6 Å. This reveals that the protein consists of a three-domain fold, including a previously-unreported compact β-propeller-TTR combination, in which an extended loop of the TTR plugs the β-propeller core. Electron density is observed for ten bound ions: six calcium, three potassium and one sodium. Low potassium ion concentrations lead to changes in tryptophan environment and exposure of two buried free cysteines located on a β-blade and in the β-propeller-plugging TTR loop. Mutating the two free cysteines to serines prevents covalent intermolecular interactions, but not aggregation, in absence of potassium ions. The potassium ion binding sites are located between the blades of the β-propeller, explaining their importance for the stability of the CRTAC1 fold. Despite varying in sequence, the three potassium ion binding sites are structurally similar and conserved features of the CRTAC protein family. These insights into the stability and structure of CRTAC1 provide a basis for further work into the function of CRTAC1 in health and disease.Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.