Phadtare Sangita, Severinov Konstantin
Department of Biochemistry, Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, NJ 08854, USA.
Nucleic Acids Res. 2005 Oct 6;33(17):5583-90. doi: 10.1093/nar/gki859. Print 2005.
Escherichia coli contains nine members of the CspA family. CspA and some of its homologues play critical role in cold acclimation of cells by acting as RNA chaperones, destabilizing nucleicacid secondary structures. Disruption of nucleic acid melting activity of CspE led to loss of its transcription antitermination activity and consequently its cold acclimation activity. To date, the melting activity of Csp proteins was studied using partially double-stranded model nucleic acids substrates forming stem-loop structures. Here, we studied the mechanism of nucleic acid melting by CspE. We show that CspE melts the stem region in two directions, that CspE-induced melting does not require the continuity of the substrate's loop region, and CspE can efficiently melt model substrates with single-stranded overhangs as short as 4 nt. We further show that preferential binding of CspE at the stem-loop junction site initiates melting; binding of additional CspE molecules that fully cover the single-stranded region of a melting substrate leads to complete melting of the stem.
大肠杆菌含有九个CspA家族成员。CspA及其一些同源物通过充当RNA伴侣，破坏核酸二级结构，在细胞的冷适应过程中发挥关键作用。CspE核酸解链活性的破坏导致其转录抗终止活性丧失，进而导致其冷适应活性丧失。迄今为止，使用形成茎环结构的部分双链模型核酸底物研究了Csp蛋白的解链活性。在此，我们研究了CspE核酸解链的机制。我们发现CspE沿两个方向解开茎区域，CspE诱导的解链不需要底物环区域的连续性，并且CspE可以有效地解开单链突出端短至4个核苷酸的模型底物。我们进一步表明，CspE在茎环连接位点的优先结合启动了解链；额外的CspE分子结合完全覆盖解链底物的单链区域，导致茎的完全解链。
