Bischoff J, Libresco S, Shia M A, Lodish H F
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142.
J Cell Biol. 1988 Apr;106(4):1067-74. doi: 10.1083/jcb.106.4.1067.
Antibody-induced degradation and chemical cross-linking experiments have been carried out to assess the nature of the interaction between the two asialoglycoprotein-receptor polypeptides, H1 and H2, synthesized in HepG2 cells. Incubation of HepG2 cell monolayers with anti-H1 antibody caused a specific and equal loss of both H1 and H2 polypeptides. The same result was obtained with anti-H2 antibody. Control serum did not affect the level of H1 or H2 not did anti-H1 or anti-H2 antibodies affect the level of the transferrin receptor. The chemical cross-linking reagent, difluorodinitrobenzene, has been used to demonstrate that H1 can be cross-linked to H2 in HepG2 cell microsomal membranes. Dimer and trimer species with apparent molecular masses of 93 and 148 kD, respectively, were readily observed upon chemical cross-linking and some dimers and trimers were immunoreactive with both anti-H1 and anti-H2 antibodies. The putative trimer, possibly two H1 and one H2 molecules, is a minimum estimate of the true size of the asialoglycoprotein receptor in intact HepG2 cell, and it is possible that larger hetero-oligomeric forms of the receptor exist. The results of both types of experiments indicate that H1 and H2 form an oligomeric complex in HepG2 cells and thus, both polypeptides constitute the human asialoglycoprotein receptor.
已开展抗体诱导降解和化学交联实验,以评估在HepG2细胞中合成的两种去唾液酸糖蛋白受体多肽H1和H2之间相互作用的性质。用抗H1抗体孵育HepG2细胞单层导致H1和H2多肽均出现特异性且等量的损失。用抗H2抗体也得到了相同的结果。对照血清不影响H1或H2的水平,抗H1或抗H2抗体也不影响转铁蛋白受体的水平。化学交联试剂二氟二硝基苯已被用于证明在HepG2细胞微粒体膜中H1可与H2交联。化学交联后很容易观察到表观分子量分别为93 kD和148 kD的二聚体和三聚体,一些二聚体和三聚体与抗H1和抗H2抗体均有免疫反应性。推测的三聚体可能由两个H1分子和一个H2分子组成,这是完整HepG2细胞中去唾液酸糖蛋白受体真实大小的最小估计,并且受体可能存在更大的异源寡聚体形式。两类实验的结果均表明H1和H2在HepG2细胞中形成寡聚体复合物,因此,这两种多肽构成人去唾液酸糖蛋白受体。