Oliver J D, Roderick H L, Llewellyn D H, High S
School of Biological Sciences, University of Manchester, Manchester, M13 9PT, United Kingdom.
Mol Biol Cell. 1999 Aug;10(8):2573-82. doi: 10.1091/mbc.10.8.2573.
ERp57 is a lumenal protein of the endoplasmic reticulum (ER) and a member of the protein disulfide isomerase (PDI) family. In contrast to archetypal PDI, ERp57 interacts specifically with newly synthesized glycoproteins. In this study we demonstrate that ERp57 forms discrete complexes with the ER lectins, calnexin and calreticulin. Specific ERp57/calreticulin complexes exist in canine pancreatic microsomes, as demonstrated by SDS-PAGE after cross-linking, and by native electrophoresis in the absence of cross-linking. After in vitro translation and import into microsomes, radiolabeled ERp57 can be cross-linked to endogenous calreticulin and calnexin while radiolabeled PDI cannot. Likewise, radiolabeled calreticulin is cross-linked to endogenous ERp57 but not PDI. Similar results were obtained in Lec23 cells, which lack the glucosidase I necessary to produce glycoprotein substrates capable of binding to calnexin and calreticulin. This observation indicates that ERp57 interacts with both of the ER lectins in the absence of their glycoprotein substrate. This result was confirmed by a specific interaction between in vitro synthesized calreticulin and ERp57 prepared in solution in the absence of other ER components. We conclude that ERp57 forms complexes with both calnexin and calreticulin and propose that it is these complexes that can specifically modulate glycoprotein folding within the ER lumen.
ERp57是内质网（ER）的一种腔内蛋白，属于蛋白质二硫键异构酶（PDI）家族成员。与典型的PDI不同，ERp57能与新合成的糖蛋白特异性相互作用。在本研究中，我们证明ERp57与内质网凝集素钙连蛋白和钙网蛋白形成离散复合物。交联后进行SDS-PAGE以及在未交联情况下进行非变性电泳均表明，犬胰腺微粒体中存在特异性的ERp57/钙网蛋白复合物。体外翻译并导入微粒体后，放射性标记的ERp57可与内源性钙网蛋白和钙连蛋白交联，而放射性标记的PDI则不能。同样，放射性标记的钙网蛋白可与内源性ERp57交联，但不能与PDI交联。在Lec23细胞中也得到了类似结果，该细胞缺乏产生能够与钙连蛋白和钙网蛋白结合的糖蛋白底物所需的葡糖苷酶I。这一观察结果表明，在没有糖蛋白底物的情况下，ERp57也能与这两种内质网凝集素相互作用。体外合成的钙网蛋白与在无其他内质网成分的溶液中制备的ERp57之间的特异性相互作用证实了这一结果。我们得出结论，ERp57与钙连蛋白和钙网蛋白均形成复合物，并推测正是这些复合物能够特异性调节内质网腔内糖蛋白的折叠。
