The Complex Energy Landscape of the Protein IscU.
The Complex Energy Landscape of the Protein IscU.
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DOI：
10.1016/j.bpj.2015.07.045
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发表时间：
2015-09-01
影响因子：
3.4
通讯作者：
Markley JL
中科院分区：
文献类型：
--
作者：
Bothe JR;Tonelli M;Ali IK;Dai Z;Frederick RO;Westler WM;Markley JL
IscU是大肠杆菌中铁-硫(Fe-S)簇生物合成的骨架蛋白，在铁-S簇的合成和转移过程中穿越了一个复杂的能量格局。我们以前的研究表明，ISCU存在两种相互转换的构象状态：一种是结构化的(S)，另一种是大体上无序的(D)。这两种状态似乎在功能上都很重要，因为参与Fe-S簇组装或转移的蛋白质已被证明优先与IscU的S或D状态相互作用。利用核磁共振谱、小角X射线散射(SAXS)和差示扫描量热法表征了ISCU的复杂结构-能量分布。结果表明，在pH 8.0时，ISCU的S态在25°C时最大，加热或冷却使蛋白质向D态转变。核磁共振谱和差示扫描量热分析结果表明，S→D的热诱导和冷诱导转变都是协同的和双态的。核磁共振和SAXS的低分辨结构信息表明，冷诱导和热诱导的D态结构相似。核磁共振结果表明，这两种状态具有相似的1H-15N HSQC谱和相同的肽基-脯氨酸键构型，并且通过小角X射线光谱分析，两者似乎都比S状态有类似的扩展。虽然在其他蛋白质中，冷变性状态被发现比热变性状态略紧，但这两种状态在ISCU中占据了相似的体积。
IscU, the scaffold protein for iron-sulfur (Fe-S) cluster biosynthesis in Escherichia coli, traverses a complex energy landscape during Fe-S cluster synthesis and transfer. Our previous studies showed that IscU populates two interconverting conformational states: one structured (S) and one largely disordered (D). Both states appear to be functionally important because proteins involved in the assembly or transfer of Fe-S clusters have been shown to interact preferentially with either the S or D state of IscU. To characterize the complex structure-energy landscape of IscU, we employed NMR spectroscopy, small-angle x-ray scattering (SAXS), and differential scanning calorimetry. Results obtained for IscU at pH 8.0 show that its S state is maximally populated at 25°C and that heating or cooling converts the protein toward the D state. Results from NMR and DSC indicate that both the heat- and cold-induced S→D transitions are cooperative and two-state. Low-resolution structural information from NMR and SAXS suggests that the structures of the cold-induced and heat-induced D states are similar. Both states exhibit similar 1H-15N HSQC spectra and the same pattern of peptidyl-prolyl peptide bond configurations by NMR, and both appear to be similarly expanded compared with the S state based on analysis of SAXS data. Whereas in other proteins the cold-denatured states have been found to be slightly more compact than the heat-denatured states, these two states occupy similar volumes in IscU.