Jaynes E N, Grant P G, Giangrande G, Wieder R, Cooperman B S
Biochemistry. 1978 Feb 21;17(4):561-9. doi: 10.1021/bi00597a001.
The photoincorporation of puromycin into Escherichia coli ribosomes has been studied in detail. Incorporation into protein L23 as a function of puromycin concentration follows a simple saturation curve and is specifically blocked by structural and functional analogues of puromycin, thus demonstrating that such incorporation proceeds via an affinity labeling process. Incorporation into L23 becomes more specific as the light fluence is reduced, indicating that such incorporation takes place from a native rather than light-denatured puromycin site. L23 remains the major labeled protein using ribosomes prepared by several procedures, suggesting the conservative nature of the site. In addition evidence is presented for affinity labeling of S14 and of a site in the RNA fraction of the 50S particle. Specific incorporation appears to proceed with an anomalously high quantum yield. The detailed photochemical mechanism is not understood, although 8-alkylation of purine moiety has been excluded. Incorporation is largely inhibited in the presence of thiol reagents.
对嘌呤霉素掺入大肠杆菌核糖体的光掺入过程进行了详细研究。嘌呤霉素掺入蛋白质L23的量随嘌呤霉素浓度的变化呈现简单的饱和曲线，并且被嘌呤霉素的结构和功能类似物特异性阻断，从而表明这种掺入是通过亲和标记过程进行的。随着光通量降低，掺入L23的特异性增强，这表明这种掺入发生在天然而非光变性的嘌呤霉素位点。使用多种方法制备的核糖体时，L23仍然是主要的标记蛋白，这表明该位点具有保守性。此外，还提供了S14和50S颗粒RNA部分中一个位点的亲和标记证据。特异性掺入似乎以异常高的量子产率进行。尽管已排除嘌呤部分的8-烷基化，但详细的光化学机制尚不清楚。在硫醇试剂存在下，掺入受到很大抑制。
