Mannose 6-phosphate-independent endocytosis of β-glucuronidase
Mannose 6-phosphate-independent endocytosis of β-glucuronidase
复制标题
β-葡萄糖醛酸酶的甘露糖 6-磷酸独立内吞作用
DOI：
10.1016/s0167-4889(00)00139-7
复制
发表时间：
2001
期刊：
影响因子：
--
通讯作者：
C. Michalak
中科院分区：
文献类型：
--
作者：
A. González;C. Michalak
从牛肝膜中分离到一种新的结合蛋白，它识别链霉蛋白酶消化的牛β-葡萄糖醛酸酶的特异性肽序列。先前的工作表明，该肽（IIIb 2）含有Ser-X-Ser序列，其中X可能是翻译后修饰的Trp。该受体从总牛肝膜中洗涤剂提取，并通过牛β-葡萄糖醛酸酶-Sepharose和IIIb 2肽-Sepharose柱上的亲和层析纯化。牛β-葡萄糖醛酸苷酶与分离的受体的结合需要二价阳离子，并且它们的存在对于维持受体-配体复合物是必要的。只有含有组分IIIb 2的肽序列能够削弱牛酶与受体的结合，来自牛β-葡萄糖醛酸糖苷酶的其他肽对结合没有影响。当在还原条件下通过SDS-PAGE分析时，观察到两条带，78 kDa的主带和72 kDa的弱带。兔抗这种结合蛋白的抗体揭示了存在于牛肝，人和牛成纤维细胞膜中的78 kDa蛋白。这些抗体损害人成纤维细胞对牛β-葡萄糖醛酸苷酶的内吞作用，但不损害人β-葡萄糖醛酸苷酶的内吞作用，后者被识别酶中磷酸甘露糖基部分的300 kDa受体摄取。
A new binding protein, which recognizes a specific peptide sequence from pronase digested bovine β-glucuronidase, has been isolated from bovine liver membranes. Prior work has shown that this peptide (IIIb2) contains a Ser–X–Ser sequence, where X might be a posttranslational modified Trp. This receptor was detergent-extracted from total bovine liver membranes and purified by affinity chromatography on a bovine β-glucuronidase–Sepharose and a IIIb2 peptide–Sepharose column. Binding of bovine β-glucuronidase to the isolated receptor requires divalent cations, and their presence was necessary to maintain the receptor–ligand complex. Only the peptide sequence containing the fraction IIIb2 was able to impair the binding of the bovine enzyme to the receptor, no other peptide from bovine β-glucuronidase had an effect on binding. When analyzed by SDS–PAGE under reducing conditions, two bands were observed, a major band of 78 kDa and a faint band of 72 kDa. Rabbit antibodies against this binding protein revealed the presence of the 78 kDa protein in membranes from bovine liver, human and bovine fibroblasts. These antibodies impaired human fibroblasts endocytosis of the bovine but not of the human β-glucuronidase, which is taken up by a 300 kDa receptor that recognizes phosphomannosyl moieties in the enzyme.