Laminin binding protein, 34/67 laminin receptor, carries stage-specific embryonic antigen-4 epitope defined by monoclonal antibody Raft.2.
Laminin binding protein, 34/67 laminin receptor, carries stage-specific embryonic antigen-4 epitope defined by monoclonal antibody Raft.2.
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DOI：
10.1016/j.bbrc.2005.05.052
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发表时间：
2005-07
影响因子：
3.1
通讯作者：
Y. Katagiri;N. Kiyokawa;Kyoko Nakamura;H. Takenouchi;T. Taguchi;H. Okita;A. Umezawa;J. Fujimoto
中科院分区：
文献类型：
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作者：
Y. Katagiri;N. Kiyokawa;Kyoko Nakamura;H. Takenouchi;T. Taguchi;H. Okita;A. Umezawa;J. Fujimoto
我们先前生产了针对洗涤剂不溶性微区的单克隆抗体，即，人肾癌细胞系ACHN的筏微结构域。Raft.2是这些单克隆抗体中的一种，其识别具有阶段特异性胚胎抗原（SSEA）-4表位的唾液酸globopentaosylceramide。尽管小鼠胚胎癌（EC）细胞系F9不表达SSEA-4，但一些F9细胞用Raft.2染色。Western分析和基质辅助激光解吸电离-飞行时间质谱鉴定了Raft.2结合分子为层粘连蛋白结合蛋白（LBP），即，34/67层粘连蛋白受体。弱酸处理或用产气荚膜梭菌唾液酸酶消化减少了Raft.2与硝酸纤维素片上的LBP的结合，并且[14 C]半乳糖掺入LBP中，表明LBP具有唾液酸化的碳水化合物部分。通过蔗糖密度梯度离心和共聚焦显微镜检查的亚细胞定位分析显示LBP被定位在质膜的外表面上。SSEA-4阳性人EC细胞系NCR-G3细胞也表达Raft.2结合LBP。
We previously produced monoclonal antibodies against the detergent-insoluble microdomain, i.e., the raft microdomain, of the human renal cancer cell line ACHN. Raft.2, one of these monoclonal antibodies, recognizes sialosyl globopentaosylceramide, which has the stage-specific embryonic antigen (SSEA)-4 epitope. Although the mouse embryonal carcinoma (EC) cell line F9 does not express SSEA-4, some F9 cells stained with Raft.2. Western analysis and matrix-assisted laser desorption ionization-time of flight mass spectrometry identified the Raft.2 binding molecule as laminin binding protein (LBP), i.e., 34/67 laminin receptor. Weak acid treatment or digestion with Clostridium perfringens sialidase reduced Raft.2 binding to LBP on nitrocellulose sheets and [14C]galactose was incorporated into LBP, indicating LBP to have a sialylated carbohydrate moiety. Subcellular localization analysis by sucrose density-gradient centrifugation and examination by confocal microscopy revealed LBP to be localized on the outer surface of the plasma membrane. An SSEA-4-positive human EC cell line, NCR-G3 cells, also expressed Raft.2-binding LBP.