Structural Basis of Substrate Specificity of Human Oligosaccharyl Transferase Subunit N33/Tusc3 and Its Role in Regulating Protein N-Glycosylation
Structural Basis of Substrate Specificity of Human Oligosaccharyl Transferase Subunit N33/Tusc3 and Its Role in Regulating Protein N-Glycosylation
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DOI：
10.1016/j.str.2014.02.013
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发表时间：
2014-04-08
期刊：
影响因子：
5.7
通讯作者：
Glockshuber, Rudi
中科院分区：
文献类型：
--
作者：
Mohorko, Elisabeth;Owen, Robin L.;Glockshuber, Rudi
蛋白质在内质网（ER）中的N-连接糖基化是真核生物必需的，并由寡糖基转移酶（OST）催化。人OST是七个亚基的异源寡聚体。亚基N33/Tusc 3是一种肿瘤抑制候选者，亚基N33/Tusc 3的缺陷与非综合征性精神发育迟滞有关。在这里，我们表明，N33/Tusc 3具有一个膜锚定的N-末端硫氧还蛋白结构域位于ER腔，可能会形成短暂的混合二硫键复合物与OST基板。N33/Tusc 3和两种不同的肽作为模型底物之间的复合物的X-射线结构揭示了一个定义的肽结合槽相邻的活性位点，可以容纳肽在相反的方向。结构和生物化学数据表明，N33/Tusc 3更喜欢带有疏水残基的肽，该疏水残基距离与N33/Tusc 3形成混合二硫化物的半胱氨酸两个残基。我们的研究结果支持了一个模型，其中N33/Tusc 3通过减缓糖蛋白折叠来增加人类糖蛋白亚组的糖基化效率。
N-linked glycosylation of proteins in the endoplasmic reticulum (ER) is essential in eukaryotes and catalyzed by oligosaccharyl transferase (OST). Human OST is a hetero-oligomer of seven subunits. The subunit N33/Tusc3 is a tumor suppressor candidate, and defects in the subunit N33/Tusc3 are linked with non-syndromic mental retardation. Here, we show that N33/Tusc3 possesses a membrane-anchored N-terminal thioredoxin domain located in the ER lumen that may form transient mixed disulfide complexes with OST substrates. X-ray structures of complexes between N33/Tusc3 and two different peptides as model substrates reveal a defined peptide-binding groove adjacent to the active site that can accommodate peptides in opposite orientations. Structural and biochemical data show that N33/Tusc3 prefers peptides bearing a hydrophobic residue two residues away from the cysteine forming the mixed disulfide with N33/Tusc3. Our results support a model in which N33/Tusc3 increases glycosylation efficiency for a subset of human glycoproteins by slowing glycoprotein folding.