BACKGROUND Olfactomedin-1 (Olfm1; also known as Noelin or Pancortin) is a highly-expressed secreted brain and retina protein and its four isoforms have different roles in nervous system development and function. Structural studies showed that the long Olfm1 isoform BMZ forms a disulfide-linked tetramer with a V-shaped architecture. The tips of the Olfm1 "V" each consist of two C-terminal β-propeller domains that enclose a calcium binding site. Functional characterisation of Olfm1 may be aided by new biochemical tools derived from these core structural elements. RESULTS Here we present the production, purification and structural analysis of three novel monomeric, dimeric and tetrameric forms of mammalian Olfm1 for functional studies. We characterise these constructs structurally by high-resolution X-ray crystallography and small-angle X-ray scattering. The crystal structure of the Olfm1 β-propeller domain (to 1.25 Å) represents the highest-resolution structure of an olfactomedin family member to date, revealing features such as a hydrophilic tunnel containing water molecules running into the core of the domain where the calcium binding site resides. The shorter Olfactomedin-1 isoform BMY is a disulfide-linked tetramer with a shape similar to the corresponding region in the longer BMZ isoform. CONCLUSIONS These recombinantly-expressed protein tools should assist future studies, for example of biophysical, electrophysiological or morphological nature, to help elucidate the functions of Olfm1 in the mature mammalian brain. The control over the oligomeric state of Olfm1 provides a firm basis to better understand the role of Olfm1 in the (trans-synaptic) tethering or avidity-mediated clustering of synaptic receptors such as post-synaptic AMPA receptors and pre-synaptic amyloid precursor protein. In addition, the variation in domain composition of these protein tools provides a means to dissect the Olfm1 regions important for receptor binding.

中文翻译：

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olfactomedin-1 变体的设计和结构表征作为功能研究的工具。

背景 Olfactomedin-1（Olfm1；也称为 Noelin 或 Pancortin）是一种高表达的分泌性脑和视网膜蛋白，其四种亚型在神经系统发育和功能中具有不同的作用。结构研究表明，长的 Olfm1 亚型 BMZ 形成具有 V 形结构的二硫键连接的四聚体。Olfm1“V”的每个尖端都包含两个包含钙结合位点的 C 端 β-螺旋桨结构域。来自这些核心结构元素的新生化工具可能有助于 Olfm1 的功能表征。结果在这里，我们展示了哺乳动物 Olfm1 的三种新型单体、二聚体和四聚体形式的生产、纯化和结构分析，用于功能研究。我们通过高分辨率 X 射线晶体学和小角度 X 射线散射在结构上表征这些构造。Olfm1 β-螺旋桨结构域（至 1.25 Å）的晶体结构代表了迄今为止嗅觉调节蛋白家族成员的最高分辨率结构，揭示了诸如含有水分子的亲水隧道等特征，这些水分子进入钙结合区域的核心站点驻留。较短的 Olfactomedin-1 亚型 BMY 是二硫键连接的四聚体，其形状类似于较长的 BMZ 亚型中的相应区域。结论 这些重组表达的蛋白质工具应该有助于未来的研究，例如生物物理学、电生理学或形态学性质，以帮助阐明 Olfm1 在成熟哺乳动物大脑中的功能。对 Olfm1 寡聚状态的控制为更好地理解 Olfm1 在突触后 AMPA 受体和突触前淀粉样前体蛋白等突触受体（跨突触）束缚或亲和力介导的聚集中的作用提供了坚实的基础。此外，这些蛋白质工具的结构域组成的变化提供了一种剖析对受体结合很重要的 Olfm1 区域的方法。