Characterization of ObgE Protein, the Escherichia.coli Homologue of Obg GTPas

我们在大肠杆菌中发现了ObgE(以前称为YhbZ)，是B.subtillis中Obg的同源物。它携带保守的GTP酶活性。我们已经证明了ObgE对大肠杆菌的生长是必不可少的，因为ObgE是成功的染色体分割所必需的。1.ObgE的工作原理尚不清楚。通过使用RFHR 2-D PAGE方法对一个obgE突变体进行蛋白质组分析，我们发现在obgE突变体不允许的条件下生长的样品上，30S核糖体蛋白的三个点S6、S18和S21的位置发生了变化。在野生型细胞中，S6和S18分别被RimK和Rimi蛋白翻译后修饰，在C-末端添加谷氨酸残基，在N-末端乙酰化丙氨酸残基。在ObgE缺陷条件下，S6蛋白的斑点向凝胶的酸性侧移动，这意味着S6蛋白的C-末端比平时增加了更多的谷氨酸残基。MALDI-TOFMS分析证实，还出现了一个新的非乙酰化的S18蛋白斑点，S21斑点的信号减弱。由于所有三种核糖体蛋白(S6、S18和S21)都与16S RNA的中心区结合，这一数据表明30S核糖体的组装受到ObgE缺陷条件的影响1)Kobayashi，G.，Moriya S.和Wada，C.(2001)Mol Microbiol，41,1037-1052。

We found ObgE (previously known as YhbZ) in Escherichia coli, is homologue of Obg in B. subtillis. It carries conserved GTPase activity. We have shown that ObgE is essential for E. coli growth because ObgE is required for successful chromosome partition^1. How ObgE works is still unclear.By proteome analysis of an obgE mutant using the RFHR 2-D PAGE method, we have found that the position of three spots of 30S ribosomal proteins, S6, S18 and S21 were changed on the sample grown under the non-permissive condition of the obgE mutant. In the wild type cell, S6 and S18 are modified post-translationally by RimK and RimI proteins with the addition of Glutamic acid residues to the C-terminals, and the acetylation of Alanine residue to the N-terminals, respectively. Under ObgE defective conditions, the spot of S6 protein shifted to the acidic side of the gel, which implies that more Glutamic acid residues were added to the C-terminals of the S6 protein than usual. Also a new spot of non-acetylated S18 protein appeared, which was confirmed by MALDI-TOFMS analysis, and the signal of the S21 spot was weakened. Furthermore, we found that the maturation of 16S RNA from pre-16S RNA was also inhibited under the ObgE defective conditions.Since all three ribosomal proteins (S6, S18 and S21) are known to bind to the central region of 16S RNA, this data suggests that the assembly of the 30S ribosome is affected by obgE defective conditions.1) Kobayashi, G., Moriya S., and Wada, C. (2001) Mol Microbiol., 41, 1037-1052.