Replacement of L7/L12.L10 Protein Complex in Escherichia coli Ribosomes with the Eukaryotic Counterpart Changes the Specificity of Elongation Factor Binding*
Replacement of L7/L12.L10 Protein Complex in Escherichia coli Ribosomes with the Eukaryotic Counterpart Changes the Specificity of Elongation Factor Binding*
复制标题
用真核对应物替换大肠杆菌核糖体中的 L7/L12.L10 蛋白复合物改变了延伸因子结合的特异性*
DOI：
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发表时间：
1999
影响因子：
4.8
通讯作者：
A. Hachimori
中科院分区：
文献类型：
--
作者：
T. Uchiumi;Kei Hori;Takao Nomura;A. Hachimori
大肠杆菌核糖体中的L 8蛋白复合物由L7/L12和L10组成，其组装在23 S rRNA的保守区域上，称为GTP酶相关结构域。我们替换了E中的L 8复合体。大肠杆菌50 S亚基与大鼠对应的P蛋白复合物组成的P1，P2，和P0。在30 °C下，用50%乙醇、10 mM MgCl 2、0.5 mM NH 4Cl从核糖体中除去L 8复合物，并将大鼠P复合物结合到核心颗粒上。通过加入覆盖E. coli 23 SrRNA与大鼠P复合物结合强烈，表明RNA结构域在此掺入中的直接作用。所得的杂交核糖体显示真核转位酶延伸因子（EF）-2依赖，但不是原核EF-G依赖，GTdR活性与大鼠80 S核糖体。EF-2依赖的活性依赖于P复合物的结合，并被抗生素硫链丝菌肽（一种原核核糖体GTP酶相关结构域的配体）抑制。该杂交系统清楚地显示了P复合物与GTP酶相关的RNA结构域的结合对于EF-2与核糖体的相互作用的重要性。结果还表明，E. coli 23 SrRNA参与了真核细胞在杂交系统中的转座酶依赖性GT3活性。
The L8 protein complex consisting of L7/L12 and L10 in Escherichia coli ribosomes is assembled on the conserved region of 23 S rRNA termed the GTPase-associated domain. We replaced the L8 complex in E. coli 50 S subunits with the rat counterpart P protein complex consisting of P1, P2, and P0. The L8 complex was removed from the ribosome with 50% ethanol, 10 mm MgCl2, 0.5 m NH4Cl, at 30 °C, and the rat P complex bound to the core particle. Binding of the P complex to the core was prevented by addition of RNA fragment covering the GTPase-associated domain of E. coli 23 S rRNA to which rat P complex bound strongly, suggesting a direct role of the RNA domain in this incorporation. The resultant hybrid ribosomes showed eukaryotic translocase elongation factor (EF)-2-dependent, but not prokaryotic EF-G-dependent, GTPase activity comparable with rat 80 S ribosomes. The EF-2-dependent activity was dependent upon the P complex binding and was inhibited by the antibiotic thiostrepton, a ligand for a portion of the GTPase-associated domain of prokaryotic ribosomes. This hybrid system clearly shows significance of binding of the P complex to the GTPase-associated RNA domain for interaction of EF-2 with the ribosome. The results also suggest that E. coli 23 S rRNA participates in the eukaryotic translocase-dependent GTPase activity in the hybrid system.
影响因子：
11.4
作者：
CZWORKOWSKI, J;WANG, J;MOORE, PB
通讯作者：
MOORE, PB
DOI：
10.1073/pnas.95.11.6134
发表时间：
1998-05-26
影响因子：
11.1
作者：
Agrawal, RK;Penczek, P;Frank, J
通讯作者：
Frank, J