Andersen Mikkel H, Berglund Lars, Petersen Torben E, Rasmussen Jan T
Protein Chemistry Laboratory, Department of Molecular and Structural Biology, University of Aarhus, Aarhus C, Denmark.
Biochem Biophys Res Commun. 2002 Mar 29;292(2):550-7. doi: 10.1006/bbrc.2002.6673.
Bovine lactadherin binds to the alpha(v)beta(3) and alpha(v)beta(5) integrins in an RGD-dependent manner and also to anionic phospholipids. During the affinity purification of lactadherin binding receptors, a 35-kDa protein persistently coeluted with the alpha(v)beta(5) integrin receptor. Subsequently, peptide mapping, amino acid sequencing, and mass spectrometry analysis identified this protein as bovine annexin-V. Annexin-V accompanied the integrin receptor eluted with either RGD peptide or with EDTA suggesting that annexin-V bound specifically to the alpha(v)beta(5) integrin. To further investigate this putative interaction of annexin-V with the alpha(v)beta(5) integrin receptor, human annexin-V and intracellular domains of the human alpha(v)beta(5) integrin subunits were used in ligand blotting assays. Radiolabeled annexin-V showed weak binding to the intracellular part of beta(5) integrin subunit. However, by adding the aminophospholipid, phosphatidyl serine, the interaction with the beta(5) cytoplasmic peptide was enhanced many fold. Furthermore, the interaction was shown to be independent of phosphorylation, as annexin-V bound to unphosphorylated beta(5) peptide at a similar level to the phosphorylated peptide. Since binding of annexin-V to the alpha(v) integrin subunit tail was not detected, annexin-V was shown to associate specifically with the beta(5) cytoplasmic tail. Together these findings suggest a novel link between annexins and the integrin receptor family.
牛乳糖粘连蛋白以RGD依赖的方式与α(v)β(3)和α(v)β(5)整合素结合，还能与阴离子磷脂结合。在乳糖粘连蛋白结合受体的亲和纯化过程中，一种35 kDa的蛋白质始终与α(v)β(5)整合素受体共洗脱。随后，肽图谱分析、氨基酸测序和质谱分析确定该蛋白质为牛膜联蛋白-V。膜联蛋白-V与用RGD肽或EDTA洗脱的整合素受体相伴，这表明膜联蛋白-V特异性结合α(v)β(5)整合素。为了进一步研究膜联蛋白-V与α(v)β(5)整合素受体的这种假定相互作用，在配体印迹分析中使用了人膜联蛋白-V和人α(v)β(5)整合素亚基的胞内结构域。放射性标记的膜联蛋白-V与β(5)整合素亚基的胞内部分显示出弱结合。然而，通过添加氨基磷脂磷脂酰丝氨酸，与β(5)胞质肽的相互作用增强了许多倍。此外，这种相互作用显示与磷酸化无关，因为膜联蛋白-V与未磷酸化的β(5)肽的结合水平与磷酸化肽相似。由于未检测到膜联蛋白-V与α(v)整合素亚基尾部的结合，表明膜联蛋白-V特异性地与β(5)胞质尾部结合。这些发现共同表明膜联蛋白与整合素受体家族之间存在新的联系。
