Neurofibromin is a tumor suppressor encoded by the NF1 gene, which is mutated in Rasopathy disease neurofibromatosis type I. Defects in NF1 lead to aberrant signaling through the RAS-mitogen-activated protein kinase pathway due to disruption of the neurofibromin GTPase-activating function on RAS family small GTPases. Very little is known about the function of most of the neurofibromin protein; to date, biochemical and structural data exist only for its GAP domain and a region containing a Sec-PH motif. To better understand the role of this large protein, here we carried out a series of biochemical and biophysical experiments, including size-exclusion chromatography-multiangle light scattering (SEC-MALS), small-angle X-ray and neutron scattering, and analytical ultracentrifugation, indicating that full-length neurofibromin forms a high-affinity dimer. We observed that neurofibromin dimerization also occurs in human cells and likely has biological and clinical implications. Analysis of purified full-length and truncated neurofibromin variants by negative-stain EM revealed the overall architecture of the dimer and predicted the potential interactions that contribute to the dimer interface. We could reconstitute structures resembling high-affinity full-length dimers by mixing N- and C-terminal protein domains in vitro The reconstituted neurofibromin was capable of GTPase activation in vitro, and co-expression of the two domains in human cells effectively recapitulated the activity of full-length neurofibromin. Taken together, these results suggest how neurofibromin dimers might form and be stabilized within the cell.

中文翻译：

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生化和结构分析表明肿瘤抑制神经纤维蛋白 (NF1) 形成高亲和力二聚体。


神经纤维蛋白是一种由 NF1 基因编码的肿瘤抑制因子，该基因在 I 型 Rasopathy 疾病神经纤维瘤病中发生突变。由于 RAS 上的神经纤维蛋白 GTP 酶激活功能受到破坏，NF1 缺陷会导致 RAS 丝裂原激活蛋白激酶通路信号异常家庭小型 GTPases。人们对大多数神经纤维蛋白的功能知之甚少。迄今为止，仅存在其 GAP 结构域和包含 Sec-PH 基序的区域的生化和结构数据。为了更好地了解这种大蛋白质的作用，我们在这里进行了一系列生化和生物物理实验，包括尺寸排阻色谱-多角度光散射（SEC-MALS）、小角度X射线和中子散射以及分析超速离心，表明全长神经纤维蛋白形成高亲和力二聚体。我们观察到神经纤维蛋白二聚化也发生在人类细胞中，并且可能具有生物学和临床意义。通过负染色电镜对纯化的全长和截短的神经纤维蛋白变体进行分析，揭示了二聚体的整体结构，并预测了有助于二聚体界面的潜在相互作用。我们可以通过在体外混合 N 端和 C 端蛋白结构域来重建类似于高亲和力全长二聚体的结构。重建的神经纤维蛋白能够在体外激活 GTP 酶，并且这两个结构域在人体细胞中的共表达有效地重现了活性全长神经纤维蛋白。总而言之，这些结果表明神经纤维蛋白二聚体如何在细胞内形成并稳定。