Kirillov S V, Makhno V I, Semenkov Y P
Nucleic Acids Res. 1980 Jan 11;8(1):183-96. doi: 10.1093/nar/8.1.183.
30S subunits were isolated capable to bind simultaneously two molecules of Phe-tRNAPhe (or N-Acetyl-Phe-tRNAPhe), both poly(U) dependent. The site with higher affinity to tRNA was identified as P site. tRNA binding to this site was not inhibited by low concentrations of tetracycline (2 x 10(-5)M) and, on the other hand, N-Acetyl-Phe-tRNAPhe, initially prebound to the 30S.poly(U) complex in the presence of tetracycline, reacted with puromycin quantitatively after addition of 50S subunits. The site with lower affinity to tRNA revealed features of the A site: tetracycline fully inhibited the binding of both Phe-tRNAPhe and N-Acetyl-Phe-tRNAPhe. Binding of two molecules of Phe-tRNAPhe to the 30S.poly(U) complex followed by the addition of 50S subunits resulted in the formation of (Phe)2-tRNAPhe in 75-90% of the reassociated 70S ribosomes. These results prove that isolated 30S subunits contain two physically distinct centers for the binding of specific aminoacyl- (or peptidyl-) tRNA. Addition of 50S subunits results in the formation of whole 70S ribosomes with usual donor and acceptor sites.
分离出的30S亚基能够同时结合两个苯丙氨酰 - tRNAphe（或N - 乙酰 - 苯丙氨酰 - tRNAphe）分子，二者均依赖于聚尿苷酸。对tRNA具有较高亲和力的位点被鉴定为P位点。低浓度的四环素（2×10⁻⁵M）不会抑制tRNA与该位点的结合，另一方面，在四环素存在下最初预结合到30S·聚尿苷酸复合物上的N - 乙酰 - 苯丙氨酰 - tRNAphe，在加入50S亚基后与嘌呤霉素发生定量反应。对tRNA具有较低亲和力的位点显示出A位点的特征：四环素完全抑制苯丙氨酰 - tRNAphe和N - 乙酰 - 苯丙氨酰 - tRNAphe的结合。两个苯丙氨酰 - tRNAphe分子与30S·聚尿苷酸复合物结合，随后加入50S亚基，在75 - 90%重新结合的70S核糖体中形成了（苯丙氨酸）₂ - tRNAphe。这些结果证明，分离出的30S亚基包含两个物理上不同的特异性氨酰 - （或肽酰 - ）tRNA结合中心。加入50S亚基会导致形成具有通常供体位点和受体位点的完整70S核糖体。
