Mallet W G, Brodsky F M
Department of Biopharmaceutical Sciences, School of Pharmacy, University of California, San Francisco, USA.
J Cell Sci. 1996 Dec;109 ( Pt 13):3059-68. doi: 10.1242/jcs.109.13.3059.
Adaptors are the membrane-binding components of clathrin-coated vesicles. The interaction of the trans-Golgi coat adaptor AP1 with membrane-associated proteins was analyzed by affinity chromatography. Proteins of 83 and 52 kDa bound specifically to the core domain of AP1 and showed no interaction with AP2 or other clathrin-coated vesicle proteins. The AP1-binding proteins were tightly membrane-associated, though behaved as peripheral membrane proteins. They were detected in membranes depleted of clathrin-coated vesicles and not in coated vesicles, suggesting that the interaction of these proteins with AP1 may precede coated vesicle budding. Co-fractionation of the AP1-binding proteins with trans-Golgi network membrane was also observed. Upon gel filtration, both AP1-binding proteins eluted in a high molecular mass complex which was labile at high concentrations of Tris. The 83 kDa protein bound to AP1 affinity resin in the absence of the 52 kDa protein. In contrast, the separated 52 kDa protein did not bind AP1, suggesting that the 83 kDa protein is the AP1-binding component of the complex. Characterization of this protein complex defines a novel membrane-associated component that specifically interacts with AP1 and may contribute to its function in forming clathrin-coated vesicles.
衔接蛋白是网格蛋白包被小泡的膜结合成分。通过亲和层析分析了反式高尔基体衣被衔接蛋白AP1与膜相关蛋白的相互作用。83 kDa和52 kDa的蛋白质特异性结合到AP1的核心结构域，且不与AP2或其他网格蛋白包被小泡蛋白相互作用。AP1结合蛋白紧密地与膜相关，尽管表现为外周膜蛋白。它们在不含网格蛋白包被小泡的膜中被检测到，而在包被小泡中未被检测到，这表明这些蛋白与AP1的相互作用可能先于包被小泡出芽。还观察到AP1结合蛋白与反式高尔基体网络膜的共分离。在凝胶过滤时，两种AP1结合蛋白都以高分子量复合物形式洗脱，该复合物在高浓度Tris下不稳定。在没有52 kDa蛋白的情况下，83 kDa蛋白与AP1亲和树脂结合。相反，分离出的52 kDa蛋白不结合AP1，这表明83 kDa蛋白是该复合物的AP1结合成分。对这种蛋白复合物的表征定义了一种新的膜相关成分，它特异性地与AP1相互作用，并可能有助于其在形成网格蛋白包被小泡中的功能。
