Gailit J, Ruoslahti E
Cancer Research Center, La Jolla Cancer Research Foundation, California 92037.
J Biol Chem. 1988 Sep 15;263(26):12927-32.
The cell surface receptor for fibronectin is a heterodimeric membrane protein that recognizes an Arg-Gly-Asp sequence in fibronectin and that requires cations such as Mg2+ or Ca2+ for binding to fibronectin. The divalent cation requirements of this receptor were analyzed by measuring attachment of receptor liposomes to ligand-coated surfaces in the presence of different cations. The most striking effect observed was produced by Mn2+, which increased the binding of the receptor liposomes to fibronectin 2-3-fold over their binding in buffers containing Ca2+ and Mg2+. The binding activities of two related adhesion receptors, the vitronectin receptor and platelet GP IIb-IIIa, were supported but not enhanced by Mn2+. Two observations suggest that Mn2+ can compete with Ca2+ for the same cation-binding sites of the receptor. First, Mn2+ could still enhance fibronectin receptor binding activity even in the presence of 10-fold higher concentrations of Ca2+ or Mg2+. Second, Mn2+ inhibited the binding of radioactive Ca2+ to the alpha subunit of the receptor. The increased fibronectin receptor activity in the presence of Mn2+ appeared to be due to an increase in the affinity of the receptor for the Arg-Gly-Asp sequence because a 110-kDa cell attachment fragment and a synthetic hexapeptide containing the Arg-Gly-Asp sequence inhibited liposome binding more effectively in the presence of Mn2+ than in the presence of Ca2+/Mg2+. The affinity for the peptide was affected more than the affinity for the fragment, indicating that Mn2+ also induces a change in receptor specificity. Increased receptor binding in the presence of Mn2+ was also apparent in affinity chromatography of the fibronectin receptor on the 110-kDa fibronectin fragment; Mn2+ improved the yield of the receptor 4-fold. Mn2+ similarly increased the number of receptor-fibronectin complexes in preparations analyzed by electron microscopy. These results show that exogenous influences can modulate the affinity and specificity with which the fibronectin receptor binds to its ligands.
纤连蛋白的细胞表面受体是一种异源二聚体膜蛋白，它能识别纤连蛋白中的精氨酸 - 甘氨酸 - 天冬氨酸序列，并且需要镁离子（Mg2+）或钙离子（Ca2+）等阳离子才能与纤连蛋白结合。通过在不同阳离子存在的情况下测量受体脂质体与配体包被表面的附着情况，分析了该受体对二价阳离子的需求。观察到的最显著效应是由锰离子（Mn2+）产生的，与在含有钙离子和镁离子的缓冲液中相比，锰离子使受体脂质体与纤连蛋白的结合增加了2 - 3倍。玻璃粘连蛋白受体和血小板糖蛋白IIb - IIIa这两种相关黏附受体的结合活性受到锰离子的支持，但并未增强。两项观察结果表明，锰离子可以与钙离子竞争受体相同的阳离子结合位点。第一，即使存在浓度高出10倍的钙离子或镁离子，锰离子仍能增强纤连蛋白受体的结合活性。第二，锰离子抑制放射性钙离子与受体α亚基的结合。在锰离子存在的情况下，纤连蛋白受体活性的增加似乎是由于受体对精氨酸 - 甘氨酸 - 天冬氨酸序列的亲和力增加，因为在锰离子存在的情况下，一个110 kDa的细胞附着片段和一个含有精氨酸 - 甘氨酸 - 天冬氨酸序列的合成六肽比在钙离子/镁离子存在的情况下更有效地抑制脂质体结合。对该肽的亲和力比对片段的亲和力影响更大，这表明锰离子也会引起受体特异性的变化。在纤连蛋白受体与110 kDa纤连蛋白片段的亲和层析中，在锰离子存在的情况下受体结合的增加也很明显；锰离子使受体的产量提高了4倍。在通过电子显微镜分析的制剂中，锰离子同样增加了受体 - 纤连蛋白复合物的数量。这些结果表明，外源性影响可以调节纤连蛋白受体与其配体结合的亲和力和特异性。
