O-GlcNAc 修饰是一种保守的翻译后修饰，可将 N-乙酰氨基葡萄糖 (GlcNAc) 附着到多种细胞蛋白上。为了响应营养和激素信号，O-GlcNAcylation 通过调节靶蛋白的稳定性、结构和功能来调节多种细胞过程。 O-GlcNAc 酰化失调与癌症、糖尿病和神经退行性疾病的发病机制有关。一对酶，即 O-GlcNAc 转移酶 (OGT) 和 O-GlcNAcase (OGA)，可催化人类蛋白质组中 3,000 多种蛋白质上 O-GlcNAc 的添加和去除。然而，OGT 如何选择其天然底物并维持这么多底物针对 OGA 的 O-GlcNAc 酰化的稳态控制尚不完全清楚。在这里，我们展示了人类 OGT 和 OGT-OGA 复合物的冷冻电子显微镜 (cryo-EM) 结构。我们的研究表明，OGT 形成了一种具有重要功能的剪刀形二聚体。在OGT-OGA复合物结构中，一个长的柔性OGA片段占据了OGT延伸的底物结合沟，并定位了用于O-GlcNAcNA酰化的丝氨酸，从而防止OGT修饰其他底物。相反，OGT 破坏 OGA 的功能二聚化并封闭其活性位点，导致其他底物的进入受阻。 OGT 和 OGA 之间的这种相互抑制可能会限制无效的 O-GlcNAc 酰化循环，并有助于维持 O-GlcNAc 稳态。© 2023。作者。

O-GlcNAcylation is a conserved post-translational modification that attaches N-acetyl glucosamine (GlcNAc) to myriad cellular proteins. In response to nutritional and hormonal signals, O-GlcNAcylation regulates diverse cellular processes by modulating the stability, structure, and function of target proteins. Dysregulation of O-GlcNAcylation has been implicated in the pathogenesis of cancer, diabetes, and neurodegeneration. A single pair of enzymes, the O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), catalyzes the addition and removal of O-GlcNAc on over 3,000 proteins in the human proteome. However, how OGT selects its native substrates and maintains the homeostatic control of O-GlcNAcylation of so many substrates against OGA is not fully understood. Here, we present the cryo-electron microscopy (cryo-EM) structures of human OGT and the OGT-OGA complex. Our studies reveal that OGT forms a functionally important scissor-shaped dimer. Within the OGT-OGA complex structure, a long flexible OGA segment occupies the extended substrate-binding groove of OGT and positions a serine for O-GlcNAcylation, thus preventing OGT from modifying other substrates. Conversely, OGT disrupts the functional dimerization of OGA and occludes its active site, resulting in the blocking of access by other substrates. This mutual inhibition between OGT and OGA may limit the futile O-GlcNAcylation cycles and help to maintain O-GlcNAc homeostasis.© 2023. The Author(s).