Arai N, Kaziro Y
J Biochem. 1975 Feb;77(2):439-47. doi: 10.1093/oxfordjournals.jbchem.a130743.
At low NH4-+ concentrations, 50S ribosomal subunits from E. coli were fully active in the absence of 30S ribosomal subunits, in forming a complex with the polypeptide chain elongation factor G (EF-G) and guanine nucleotide (ternary complex formation), and also in supporting EF-G dependent hydrolysis of GTP (uncoupled GTPase reaction). However, both activities were markedly inhibited on increasing the concentration of the monovalent cation, and at 160 mM NH4-+, the optimal concentration for polypeptide synthesis in a cell-free system, almost no activity was observed with 50S ribosomes alone. It was found that the inhibitory effect of NH4-+ was reversed by addition of 30S subunits. Thus, at 160 mM NH4-+, only 70S ribosomes were active in supporting the above two EF-G dependent reactions, whereas at 20 mM NH4-+, 50S ribosomes were almost as active as 70S ribosomes. Kinetic studies on inhibition by NH4-+ of the formation of 50S ribosome-EF-G-guanine nucleotide complex, indicated that the inhibition was due to reduction in the number of active 50S ribosomes which were capable of interacting with EF-G and GTP at higher concentrations of NH4-+. The inhibitory effects of NH4-+ on ternary complex formation and the uncoupled GTPase reaction were markedly influenced by temperature, and were much greater at 0 degrees than at 30 degrees. A conformational change of 50S subunits through association with 30S subunits is suggested.
在低浓度铵离子（NH₄⁺）条件下，来自大肠杆菌的50S核糖体亚基在没有30S核糖体亚基的情况下完全具有活性，可与多肽链延伸因子G（EF-G）和鸟嘌呤核苷酸形成复合物（三元复合物形成），也能支持EF-G依赖的GTP水解（非偶联GTP酶反应）。然而，随着单价阳离子浓度的增加，这两种活性均受到显著抑制，在160 mM NH₄⁺（无细胞系统中多肽合成的最佳浓度）时，单独的50S核糖体几乎没有活性。研究发现，添加30S亚基可逆转NH₄⁺的抑制作用。因此，在160 mM NH₄⁺时，只有70S核糖体在支持上述两种EF-G依赖反应中具有活性，而在20 mM NH₄⁺时，50S核糖体的活性几乎与70S核糖体相同。对NH₄⁺抑制50S核糖体-EF-G-鸟嘌呤核苷酸复合物形成的动力学研究表明，抑制作用是由于在较高NH₄⁺浓度下能够与EF-G和GTP相互作用的活性50S核糖体数量减少所致。NH₄⁺对三元复合物形成和非偶联GTP酶反应的抑制作用受温度显著影响，在0℃时比在30℃时大得多。这表明5OS亚基通过与30S亚基结合发生了构象变化。
