Nag B, Tewari D S, Sommer A, Olson H M, Glitz D G, Traut R R
J Biol Chem. 1987 Jul 15;262(20):9681-7.
A monoclonal antibody specific for Escherichia coli ribosomal protein L5 was isolated from a cell line obtained from Dr. David Schlessinger. Its unique specificity for L5 was confirmed by one- and two-dimensional electrophoresis and immunoblotting. The antibody recognized L5 both in 50 S subunits and 70 S ribosomes. Both antibody and Fab fragments had similar effects on the ribosome functions tested. Antibody bound to 50 S subunits inhibited their reassociation with 30 S subunits at 10 mM Mg2+ but not 15 mM, the concentration present for in vitro protein synthesis. The 70 S couples were not dissociated by the antibody. The antibody caused inhibition of polyphenylalanine synthesis at molar ratios to 50 S or 70 S particles of 4:1. The major inhibitory effect was on the peptidyltransferase reaction. There was no effect on either elongation factor binding or the associated GTPase activities. The site of antibody binding to 50 S was determined by electron microscopy. Antibody was seen to bind beside the central protuberance or head of the particle, on the side away from the L7/L12 stalk, and on or near the region at which the 50 S subunit interacts with the 30 S subunit. This site of antibody binding is fully consistent with its biochemical effects.
从大卫·施莱辛格博士提供的细胞系中分离出一种对大肠杆菌核糖体蛋白L5具有特异性的单克隆抗体。通过一维及二维电泳和免疫印迹法证实了其对L5的独特特异性。该抗体在50 S亚基和70 S核糖体中均能识别L5。抗体和Fab片段对所测试的核糖体功能具有相似的作用。与50 S亚基结合的抗体在10 mM Mg2+浓度下可抑制其与30 S亚基的重新结合，但在15 mM（体外蛋白质合成时的浓度）时则无此作用。70 S核糖体对不会被该抗体解离。当抗体与50 S或70 S颗粒的摩尔比为4:1时，会抑制聚苯丙氨酸的合成。主要抑制作用发生在肽基转移酶反应上。对延伸因子结合或相关的GTP酶活性均无影响。通过电子显微镜确定了抗体与50 S的结合位点。可以看到抗体结合在颗粒的中央突起或头部旁边，位于远离L7/L12柄的一侧，以及50 S亚基与30 S亚基相互作用区域之上或附近。抗体的这个结合位点与其生化效应完全一致。
