Vince R, Brownell J, Fong K L
Biochemistry. 1978 Dec 12;17(25):5489-93. doi: 10.1021/bi00618a025.
A photoaffinity labeling puromycin analogue, Nepsilon-(2-nitro-4-azidophenyl)-L-lysinyl puromycin aminonucleoside (NAP-Lys-Pan), was synthesized and used for investigation of the peptidyl transferase center of 70S riobsomes. Visible light irradiation of NAP-Lys-Pan led to covalent linkage of the analogue with Escherichia coli ribosomes. In a subsequent step, poly(uridylic acid) was employed to direct Ac[14C]Phe-tRNA to the P sites of the photolabeled ribosomes. Transpeptidation of Ac[14C]phenylalanine to the bound NAP-Lys-Pan resulted in selective incorporation of radioactive label into the peptidyl transferase A site. Dissociation of the ribosomes into subunits, and digestion of the RNA components, indicated that the radioactive label was incorporated into a protein fraction of the 50S subunit.
合成了一种光亲和标记嘌呤霉素类似物Nε-(2-硝基-4-叠氮苯基)-L-赖氨酰嘌呤霉素氨基核苷（NAP-Lys-Pan），并用于研究70S核糖体的肽基转移酶中心。对NAP-Lys-Pan进行可见光照射会导致该类似物与大肠杆菌核糖体发生共价连接。在后续步骤中，使用聚尿苷酸将Ac[14C]苯丙氨酰-tRNA引导至光标记核糖体的P位点。将Ac[14C]苯丙氨酸转肽至结合的NAP-Lys-Pan上，导致放射性标记选择性地掺入肽基转移酶A位点。核糖体解离成亚基，并对RNA成分进行消化，结果表明放射性标记掺入了50S亚基的蛋白质组分中。
