Dobrila L, Serban G, Heltianu C
Institute of Cellular Biology and Pathology, Bucharest, Romania.
Biosci Rep. 1996 Oct;16(5):425-38. doi: 10.1007/BF01207267.
In the present work we examined whether the interaction between albumin molecules and thymocytes involves albumin-binding proteins (ABP). Two plasmalemma-rich fractions obtained by differential centrifugation from rat thymus lymphocytes were characterized biochemically and morphologically. These fractions were examined by ligand-blotting and ligand affinity chromatography techniques. Plasmalemma proteins separated by SDS-PAGE were electrotransferred onto nitrocellulose membranes and incubated with 125I-albumin, in the presence or absence of excess native albumin. The autoradiogram revealed specific binding to two sets of polypeptides of 16-18 and 29-31 kDa, which could be blocked by native albumin. To elucidate whether albumin-binding proteins are exposed on the cell surface, intact lymphocytes were surface radioiodinated and membrane fractions prepared from them were subjected to affinity chromatography on albumin-agarose beads. The protein thus purified had, like ABP, M(r) of 16 and 31. These data indicate that ABP (i) are components of thymocyte plasma membrane, (ii) have apparent molecular mass of 16-18 and 29-31 kDa, and (iii) are exposed on the outer membrane surface.
在本研究中，我们检测了白蛋白分子与胸腺细胞之间的相互作用是否涉及白蛋白结合蛋白（ABP）。通过差速离心从大鼠胸腺淋巴细胞中获得了两个富含质膜的组分，并对其进行了生化和形态学表征。采用配体印迹法和配体亲和层析技术对这些组分进行了检测。通过SDS-PAGE分离的质膜蛋白被电转移到硝酸纤维素膜上，并在有或没有过量天然白蛋白存在的情况下与125I-白蛋白一起孵育。放射自显影片显示与两组分子量分别为16 - 18 kDa和29 - 31 kDa的多肽有特异性结合，这种结合可被天然白蛋白阻断。为了阐明白蛋白结合蛋白是否暴露在细胞表面，对完整的淋巴细胞进行表面放射性碘化，然后从它们制备的膜组分在白蛋白-琼脂糖珠上进行亲和层析。如此纯化得到的蛋白质，与ABP一样，分子量为16和31。这些数据表明，ABP（i）是胸腺细胞质膜的成分，（ii）表观分子量为16 - 18 kDa和29 - 31 kDa，（iii）暴露在外膜表面。
