Conserved Regions 4.1 and 4.2 of ς70Constitute the Recognition Sites for the Anti-ς Factor AsiA, and AsiA Is a Dimer Free in Solution*
Conserved Regions 4.1 and 4.2 of ς70Constitute the Recognition Sites for the Anti-ς Factor AsiA, and AsiA Is a Dimer Free in Solution*
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DOI：
10.1074/jbc.m106400200
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发表时间：
2001-11
期刊：
影响因子：
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通讯作者：
J. Urbauer;K. Adelman;R. Urbauer;M. Simeonov;J. Gilmore;M. Żółkiewski;E. Brody
中科院分区：
文献类型：
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作者：
J. Urbauer;K. Adelman;R. Urbauer;M. Simeonov;J. Gilmore;M. Żółkiewski;E. Brody
噬菌体T4编码的AsiA蛋白与大肠杆菌RNA聚合酶的Asi 70亚基的结合是控制T4基因组转录的主要事件之一。分析超离心和NMR研究表明，游离AsiA是一个对称的二聚体，二聚体可以交换亚基。利用核磁共振技术，对AsiA和Asia 70的相互识别位点进行了解析。AsiA的N-末端一半的残基直接或间接地参与与AsiA 70的结合，而AsiA 70的两个高度保守的C-末端区域，表示为4.1和4.2，构成了整个AsiA结合结构域。对应于这些区域的肽单独且同时与AsiA紧密结合。同时结合促进AsiA中的结构变化，其模拟与AsiA的完整结合决定簇的相互作用。此外，结果表明二聚体的显着重排伴随着肽结合。因此，两个保守区域4.1和4.2都密切参与了AsiA的识别。AsiA与4.1的相互作用提供了DNA和AsiA与游离AsiA 70结合的差异能力的潜在解释，以及依赖于与AsiA 70上的单个位点结合的AsiA功能模型的机制替代。
The association of the bacteriophage T4-encoded AsiA protein with the ς70 subunit of theEscherichia coli RNA polymerase is one of the principal events governing transcription of the T4 genome. Analytical ultracentrifugation and NMR studies indicate that free AsiA is a symmetric dimer and the dimers can exchange subunits. Using NMR, the mutual recognition sites on AsiA and ς70 have been elucidated. Residues throughout the N-terminal half of AsiA are involved either directly or indirectly in binding to ς70whereas the two highly conserved C-terminal regions of ς70, denoted 4.1 and 4.2, constitute the entire AsiA binding domain. Peptides corresponding to these regions bind tightly to AsiA individually and simultaneously. Simultaneous binding promotes structural changes in AsiA that mimic interaction with the complete AsiA binding determinant of ς70. Moreover, the results suggest that a significant rearrangement of the dimer accompanies peptide binding. Thus, both conserved regions 4.1 and 4.2 are intimately involved in recognition of AsiA by ς70. The interaction of AsiA with 4.1 provides a potential explanation of the differential abilities of DNA and AsiA to bind to free ς70 and a mechanistic alternative to models of AsiA function that rely on binding to a single site on ς70.