Balaji Boovaraghan, Gilson Michele, Roy Harry
Biology Department, Rensselaer Polytechnic Institute, Troy, New York 12180-3590, USA.
Photosynth Res. 2006 Jul;89(1):43-8. doi: 10.1007/s11120-006-9067-7. Epub 2006 Jun 9.
Radioactive amino acids, when added to isolated pea chloroplasts or chloroplast extracts engaged in protein synthesis, are incorporated into Rubisco large subunits that co-migrate with native Rubisco during nondenaturing electrophoresis. We have added the transition state analog 2'-carboxyarabinitol bisphosphate (CABP) to chloroplast extracts after in organello or in vitro incorporation of radioactive amino acids into Rubisco large subunits. Upon addition of CABP the radioactive bands co-migrating with native Rubisco undergo a readily detected shift in electrophoretic mobility just as the native enzyme, thus demonstrating the ability of the newly assembled molecules to interact with this transition state analog.
当将放射性氨基酸添加到参与蛋白质合成的离体豌豆叶绿体或叶绿体提取物中时，它们会被整合到核酮糖-1,5-二磷酸羧化酶/加氧酶（Rubisco）的大亚基中，这些大亚基在非变性电泳过程中与天然Rubisco共同迁移。在放射性氨基酸在叶绿体中或体外被整合到Rubisco大亚基之后，我们向叶绿体提取物中添加了过渡态类似物2'-羧基阿拉伯糖醇双磷酸（CABP）。加入CABP后，与天然Rubisco共同迁移的放射性条带在电泳迁移率上发生了易于检测到的变化，就像天然酶一样，从而证明了新组装的分子与这种过渡态类似物相互作用的能力。
