Coupled release of eukaryotic translation initiation factors 5B and 1A from 80S ribosomes following subunit joining
Coupled release of eukaryotic translation initiation factors 5B and 1A from 80S ribosomes following subunit joining
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DOI：
10.1128/mcb.02254-06
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发表时间：
2007-03-01
影响因子：
5.3
通讯作者：
Dever, Thomas E.
中科院分区：
文献类型：
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作者：
Fringer, Jeanne M.;Acker, Michael G.;Dever, Thomas E.
翻译起始GT3真核生物翻译起始因子513（eIF 5 B）与eIFIA因子结合并在体外催化核糖体亚基连接。我们发现，在酿酒酵母中eIF 5 B的快速消耗导致eIFIA和mRNA在体内40 S亚基上的积累，与亚基连接缺陷一致。用Ala取代eIF 1A（eIFIA-5A）中的最后5个残基会损害eIF 5 B与细胞提取物中的eIF 1A和体内的40 S复合物的结合。一致地，eIF 5 B的过表达抑制了表达eIF 1A-5A的酵母中的生长和翻译起始缺陷，表明eIF 1A有助于在亚基连接之前将eIF 5 B募集到40 S亚基。GTP酶缺陷型eIF 5 B-T439 A突变体在体内积累在80 S复合物上，并与eIF 1 A一起沿着保留在体外形成的80 S复合物上。同样地，eIF 5 B和eIFIA保持与在不可水解的GDPNP存在下形成的80 S复合物相关联，而这些因子在含有GTP的测定中从80 S复合物中释放。我们认为eIFIA促进eIF 5 B与40 S亚基的结合以促进亚基连接。80 S复合物形成后，eIF 5 B的GTP水解使eIF 5 B和eIF 1A同时释放，核糖体进入蛋白质合成的延伸阶段。
The translation initiation GTPase eukaryotic translation initiation factor 513 (eIF5B) binds to the factor eIFIA and catalyzes ribosomal subunit joining in vitro. We show that rapid depletion of eIF5B in Saccharomyces cerevisiae results in the accumulation of eIFIA and mRNA on 40S subunits in vivo, consistent with a defect in subunit joining. Substituting Ala for the last five residues in eIF1A (eIFIA-5A) impairs eIF5B binding to eIF1A in cell extracts and to 40S complexes in vivo. Consistently, overexpression of eIF5B suppresses the growth and translation initiation defects in yeast expressing eIF1A-5A, indicating that eIF1A helps recruit eIF5B to the 40S subunit prior to subunit joining. The GTPase-deficient eIF5B-T439A mutant accumulated on 80S complexes in vivo and was retained along with eIF1A on 80S complexes formed in vitro. Likewise, eIF5B and eIFIA remained associated with 80S complexes formed in the presence of nonhydrolyzable GDPNP, whereas these factors were released from the 80S complexes in assays containing GTP. We propose that eIFIA facilitates the binding of eIF5B to the 40S subunit to promote subunit joining. Following 80S complex formation, GTP hydrolysis by eIF5B enables the release of both eIF5B and eIF1A, and the ribosome enters the elongation phase of protein synthesis.