结核分枝杆菌
圆二色性
核糖体
动态光散射
大肠杆菌
化学
核磁共振波谱
产量(工程)
分子质量
翻译(生物学)
单体
生物化学
核糖核酸
肺结核
材料科学
立体化学
纳米技术
酶
纳米颗粒
信使核糖核酸
病理
基因
有机化学
医学
冶金
聚合物
作者
Juanjuan Yang,Yindi Liu,Shuli Xu,Haiying Lin,Chun Meng,Donghai Lin
标识
DOI:10.1016/j.pep.2018.05.014
摘要
The trans-translation system is recognized as an excellent target for developing new drugs to rapidly sterilize Mycobacterium tuberculosis (TB) infection and significantly shorten TB treatment duration. As a vital component of the trans-translation system for rescuing stalled ribosomes, the SmpB protein from Mycobacterium tuberculosis (MtbSmpB, 1-160 a. a.) mediates tmRNA binding to stalled ribosomes through forming a complex with tmRNA. So far, few works have been conducted to prepare, characterize biophysical properties and determine three-dimensional structure for the full-length MtbSmpB protein. In the present work, we successfully expressed and purified the His-tagged full-length MtbSmpB protein in Escherichia coli with a yield of 26.9 mg from 1 L of Luria Bertani medium. We also obtained MtbSmpB with a yield of 18.5 mg from 1 L of M9 minimal medium. The MtbSmpB protein showed a single band in SDS-PAGE with a molecular weight of ∼20 kDa consistent with the measurement from MALDI-TOF-mass spectrometry. The dynamic light scattering experiment indicated that MtbSmpB existed in a monomeric form. Moreover, both circular dichroism and nuclear magnetic resonance (NMR) experiments exhibited that MtbSmpB was well structured, suggesting that it could be feasible to determine its solution structure by NMR spectroscopy. NMR titration experiments showed that MtbSmpB specifically bound to tmRNA. This work lays the essential basis for further determining the solution structure and dynamics of the full-length MtbSmpB protein.
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