State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, University of CAS, Chinese Academy of Sciences (CAS), 320 Yueyang Road, 200030, Shanghai, China.
State Key Laboratory of Biotherapy and Cancer Center, Department of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan University, 610044, Chengdu, China.
Nat Commun. 2022 May 12;13(1):2617. doi: 10.1038/s41467-022-30250-6.
Eukaryotic cells are coated with an abundance of glycosylphosphatidylinositol anchor proteins (GPI-APs) that play crucial roles in fertilization, neurogenesis, and immunity. The removal of a hydrophobic signal peptide and covalent attachment of GPI at the new carboxyl terminus are catalyzed by an endoplasmic reticulum membrane GPI transamidase complex (GPI-T) conserved among all eukaryotes. Here, we report the cryo-electron microscopy (cryo-EM) structure of the human GPI-T at a global 2.53-Å resolution, revealing an equimolar heteropentameric assembly. Structure-based mutagenesis suggests a legumain-like mechanism for the recognition and cleavage of proprotein substrates, and an endogenous GPI in the structure defines a composite cavity for the lipid substrate. This elongated active site, stemming from the membrane and spanning an additional ~22-Å space toward the catalytic dyad, is structurally suited for both substrates which feature an amphipathic pattern that matches this geometry. Our work presents an important step towards the mechanistic understanding of GPI-AP biosynthesis.
真核细胞表面覆盖着大量糖基磷脂酰肌醇锚定蛋白(GPI-APs),它们在受精、神经发生和免疫中发挥着关键作用。内质网膜 GPI 转酰胺酶复合物(GPI-T)催化去除疏水性信号肽并在新的羧基末端共价连接 GPI,该复合物在所有真核生物中都保守。在这里,我们报道了人类 GPI-T 的冷冻电镜(cryo-EM)结构,分辨率为 2.53 Å,揭示了等摩尔异五聚体组装。基于结构的突变分析表明,GPI-T 对前蛋白底物的识别和切割具有类似于莱姆蛋白酶的机制,并且结构中的内源性 GPI 定义了脂质底物的复合腔。这个拉长的活性位点源自膜并向催化二联体延伸额外的~22 Å 空间,结构上适合具有与该几何形状匹配的两亲性模式的两种底物。我们的工作朝着理解 GPI-AP 生物合成的机制迈出了重要的一步。