Evolution of Ribosomal Protein S14 Demonstrated by the Reconstruction of Chimeric Ribosomes in Bacillus subtilis
Evolution of Ribosomal Protein S14 Demonstrated by the Reconstruction of Chimeric Ribosomes in Bacillus subtilis
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DOI：
10.1128/jb.00599-20
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发表时间：
2021-05-01
影响因子：
3.2
通讯作者：
Kato-Yamada, Yasuyuki
中科院分区：
文献类型：
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作者：
Akanuma, Genki;Kawamura, Fujio;Kato-Yamada, Yasuyuki
核糖体蛋白S14可分为三种类型。第一，C+型具有Zn 2+结合基序并且是祖先的。第二种和第三种是C-短型和C-长型，这两种类型都不含Zn 2+结合基序，并且它们是Ca。长度分别为90个残基和100个残基。在本研究中，来自枯草杆菌核糖体（S14 BsC+）的C+型S14完全被来自大肠杆菌（S14 Ec）或细长聚球藻（S14 Se）的异源C-长型S14取代。令人惊讶的是，在B中，S14 Ec和S14 Se完全并入70 S核糖体中。枯草芽孢杆菌然而，携带异源S14的突变体的生长速率以及产孢效率显著降低。在这些突变体中，多核糖体组分减少，30 S和50 S亚基异常积累，表明这些突变体的细胞翻译活性降低。体外分析表明，从这些突变体中纯化的70 S核糖体组分的翻译活性降低。在这些突变体的30 S组分中，核糖体蛋白S2和S3的丰度降低，而S14的丰度没有显著降低。似乎异源S14的结合改变了30 S亚基的结构，这导致位于S14结合位点附近的S2和S3的组装效率降低。此外，我们发现S.细长体不能在B中起作用。重要信息S14是一种必需的核糖体蛋白，可能已经进化为通过用锌非依赖性序列替换锌结合基序来使细菌适应锌限制的环境。预期细菌核糖体将耐受S14的替换，因为先前预测C型S14的传播涉及水平基因转移。在本研究中，我们完全替换了B中S14的C+类型。枯草杆菌核糖体与异源长C型S14的嵌合体，并表征所得嵌合核糖体。我们的研究结果表明，B。枯草杆菌核糖体对S14的替换是允许的，但可能需要S3的协同进化来更有效地利用S14的C-长型。
Ribosomal protein S14 can be classified into three types. The first, the C+ type has a Zn2+ binding motif and is ancestral. The second and third are the C- short and C- long types, neither of which contain a Zn2+ binding motif and which are ca. 90 residues and 100 residues in length, respectively. In the present study, the C+ type S14 from Bacillus subtilis ribosomes (S14BsC+) were completely replaced by the heterologous C- long type of S14 from Escherichia coli (S14Ec) or Synechococcus elongatus (S14Se). Surprisingly, S14Ec and S14Se were incorporated fully into 70S ribosomes in B. subtilis. However, the growth rates as well as the sporulation efficiency of the mutants harboring heterologous S14 were significantly decreased. In these mutants, the polysome fraction was decreased and the 30S and 50S subunits accumulated unusually, indicating that cellular translational activity of these mutants was decreased. In vitro analysis showed a reduction in the translational activity of the 70S ribosome fraction purified from these mutants. The abundance of ribosomal proteins S2 and S3 in the 30S fraction in these mutants was reduced while that of S14 was not significantly decreased. It seems likely that binding of heterologous S14 changes the structure of the 30S subunit, which causes a decrease in the assembly efficiency of S2 and S3, which are located near the binding site of S14. Moreover, we found that S3 from S. elongatus cannot function in B. subtilis unless S14Se is present.IMPORTANCE S14, an essential ribosomal protein, may have evolved to adapt bacteria to zinc-limited environments by replacement of a zinc-binding motif with a zinc-independent sequence. It was expected that the bacterial ribosome would be tolerant to replacement of S14 because of the previous prediction that the spread of C- type S14 involved horizontal gene transfer. In this study, we completely replaced the C+ type of S14 in B. subtilis ribosome with the heterologous C- long type of S14 and characterized the resulting chimeric ribosomes. Our results suggest that the B. subtilis ribosome is permissive for the replacement of S14, but coevolution of S3 might be required to utilize the C- long type of S14 more effectively.