T-rich DNA single strands bind to a preformed site on the bacterial cold shock protein Bs-CspB
T-rich DNA single strands bind to a preformed site on the bacterial cold shock protein Bs-CspB
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DOI：
10.1016/j.jmb.2006.05.044
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发表时间：
2006-07-14
影响因子：
5.6
通讯作者：
Heinemann, Udo
中科院分区：
文献类型：
--
作者：
Max, Klaas E. A.;Zeeb, Markus;Heinemann, Udo
细菌冷休克蛋白（CSPs）参与细胞对冷应激的适应。它们与单链核酸结合，KD值在微到纳摩尔范围内。在这里，我们以1.78埃的分辨率展示了枯草芽孢杆菌CspB （Bs-CspB）与六嘧啶（dT(6)）配合物的结构。Bs-CspB通过蛋白表面的两亲界面以纳米摩尔亲和力与dT6结合。单个结合亚位点通过叠加相互作用和氢键与单个核碱基相互作用。糖-磷酸主链和胸腺嘧啶核碱基的甲基仍然暴露在溶剂中，不与蛋白质基团接触。荧光滴定实验监测低聚嘧啶与Bs-CspB的结合，揭示了单个亚位点的结合偏好，并允许设计优化的七嘧啶配体，其结合具有亚纳摩尔亲和力。本研究揭示了单链核酸与Bs-CspB预形成位点结合的化学计量学和序列决定因素，从而为CSP的RNA伴侣和转录抗终止活性提供了结构基础。(c) 2006 Elsevier Ltd.版权所有。
Bacterial cold shock proteins (CSPs) are involved in cellular adaptation to cold stress. They bind to single-stranded nucleic acids with a KD value in the micro- to nanomolar range. Here we present the structure of the Bacillus subtilis CspB (Bs-CspB) in complex with hexathymidine (dT(6)) at a resolution of 1.78 angstrom. Bs-CspB binds to dT6 with nanomolar affinity via an amphipathic interface on the protein surface. Individual binding subsites interact with single nucleobases through stacking interactions and hydrogen bonding. The sugar-phosphate backbone and the methyl groups of the thymine nucleobases remain solvent exposed and are not contacted by protein groups. Fluorescence titration experiments monitoring the binding of oligopyrimidines to Bs-CspB reveal binding preferences at individual subsites and allow the design of an optimised heptapyrimidine ligand, which is bound with sub-nanomolar affinity. This study reveals the stoichiometry and sequence determinants of the binding of single-stranded nucleic acids to a preformed site on Bs-CspB and thus provides the structural basis of the RNA chaperone and transcription antitermination activities of the CSP. (c) 2006 Elsevier Ltd. All rights reserved.