A SYNTHETIC PEPTIDE DERIVED FROM THE CARBOXY TERMINUS OF THE LAMININ A-CHAIN REPRESENTS A BINDING-SITE FOR THE ALPHA-3-BETA-1-INTEGRIN
A SYNTHETIC PEPTIDE DERIVED FROM THE CARBOXY TERMINUS OF THE LAMININ A-CHAIN REPRESENTS A BINDING-SITE FOR THE ALPHA-3-BETA-1-INTEGRIN
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DOI：
10.1083/jcb.117.2.449
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发表时间：
1992-04-01
影响因子：
7.8
通讯作者：
SKUBITZ, APN
中科院分区：
文献类型：
--
作者：
GEHLSEN, KR;SRIRAMARAO, P;SKUBITZ, APN
本研究的目的是鉴定层粘连蛋白内α-3-β-1整联蛋白受体的结合位点。先前已经显示，使用蛋白水解片段和抗层粘连蛋白抗体，层粘连蛋白中用于α-3-β-1整联蛋白结合的区域定位于长臂末端的羧基末端区域（Gehlsen，K. R.，E. Engvall，K. Dickerson，W. S. Argraves和E. Ruoslahti。1989. J.Biol.Chem.264：19034-19038; Tomaselli，K. J.，D. E.霍尔湖，澳-地T.赖哈特湖A.弗里尔，K。R. Gehlsen，D. C. Turner和S.卡本内托1990. Neuron. 5：651-662）。使用合成肽，我们已经确定了层粘连蛋白A链的羧基末端区域内的氨基酸序列，其被α-3-β-1整联蛋白识别。由鼠A链的球状结构域的合成肽GD-6（编号为3011至3032的KQNCLSSRASFRGCVRNLRLSR残基）表示的氨基酸序列支持细胞附着并抑制细胞粘附于层粘连蛋白包被的表面。通过亲和层析，肽GD-6-Sepharose以阳离子依赖性方式特异性结合来自表面碘化细胞提取物的溶解的α-3-β-1，而它不结合其他整合素。此外，外源肽GD-6特异性地从层粘连蛋白-琼脂糖柱洗脱结合的α-3-β-1，但不从纤连蛋白-琼脂糖柱洗脱α-5-β-1整联蛋白。使用整联蛋白亚基特异性单克隆抗体，只有那些针对α-3和β-1亚基的抗体抑制细胞粘附到肽GD-6包被的表面。最后，针对肽GD-6的多克隆抗体与鼠和人层粘连蛋白特异性反应，并显着抑制细胞粘附到层粘连蛋白包被的表面，但不与其他基质蛋白包被的表面。这些结果将肽GD-6的层粘连蛋白A链氨基酸序列鉴定为代表层粘连蛋白中α-3-β-1整联蛋白的结合位点。
The purpose of this study was to identify the binding site(s) within laminin for the alpha-3-beta-1 integrin receptor. It has been previously shown, using proteolytic fragments and anti-laminin antibodies, that the region in laminin for alpha-3-beta-1 integrin binding is localized to the carboxy-terminal region at the end of the long arm (Gehlsen, K. R., E. Engvall, K. Dickerson, W. S. Argraves, and E. Ruoslahti. 1989. J. Biol. Chem. 264:19034-19038; Tomaselli, K. J., D. E. Hall, L. T. Reichardt, L. A. Flier, K. R. Gehlsen, D. C. Turner, and S. Carbonetto. 1990. Neuron. 5:651-662). Using synthetic peptides, we have identified an amino acid sequence within the carboxy-terminal region of the laminin A chain that is recognized by the alpha-3-beta-1 integrin. The amino acid sequence represented by the synthetic peptide GD-6 (KQNCLSSRASFRGCVRNLRLSR residues numbered 3011 to 3032) of the globular domain of the murine A chain supports cell attachment and inhibits cell adhesion to laminin-coated surfaces. By affinity chromatography, peptide GD-6-Sepharose specifically bound solubilized alpha-3-beta-1 from extracts of surface-iodinated cells in a cation-dependent manner, while it did not bind other integrins. In addition, exogenous peptide GD-6 specifically eluted bound alpha-3-beta-1 from laminin-Sepharose columns but did not elute the alpha-5-beta-1 integrin from a fibronectin-Sepharose column. Using integrin subunit-specific monoclonal antibodies, only those antibodies against the alpha-3 and beta-1 subunits inhibited cell adhesion to peptide GD-6-coated surfaces. Finally, a polyclonal antibody made against peptide GD-6 reacted specifically with both murine and human laminin and significantly inhibited cell adhesion to laminin-coated surfaces but not those coated with other matrix proteins. These results identify the laminin A chain amino acid sequence of peptide GD-6 as representing a binding site in laminin for the alpha-3-beta-1 integrin.