Monitoring the kinetics of ion-dependent protein folding by time-resolved NMR spectroscopy at atomic resolution
Monitoring the kinetics of ion-dependent protein folding by time-resolved NMR spectroscopy at atomic resolution
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DOI：
10.1021/ja994212b
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发表时间：
2000-07-05
影响因子：
15
通讯作者：
Schwalbe, H
中科院分区：
文献类型：
--
作者：
Kühn, T;Schwalbe, H
用时间分辨核磁共振波谱法监测了蛋白质再折叠的动力学。结果表明，光不稳定离子螯合剂光解可诱导金属结合蛋白的再折叠，随后Ca2+离子的释放可诱导蛋白质折叠，并且可以通过时间分辨核磁共振波谱监测共振位置的变化。该方法的可行性通过表征α -乳清蛋白（或含有Ca2+结合位点的蛋白质）在pH为7、无Ca2+的情况下在4 M尿素中展开的再折叠来证明。在300 K下，蛋白质核心残基Leu15和Val21的甲基的再折叠动力学为单指数，速率分别为0.28 s（-1）和0.23 s（-1）。
The kinetics of protein refolding have been monitored by time-resolved NMR spectroscopy. It is shown that refolding of metal binding proteins can be induced by photolysis of photo labile ion chelators, the subsequent release of Ca2+ ions can induce protein folding, and the changes in resonance positions can be monitored by time-resolved NMR spectroscopy. The feasibility of the approach is demonstrated by characterizing the refolding of alpha-lactalbumin, or protein containing a Ca2+ binding site, unfolded in 4 M urea at pH 7 in the absence of Ca2+. The refolding kinetics of the methyl groups of residues Leu15 and Val21 in the core of the protein have been determined to be mono-exponential with rates of 0.28 s(-1) and 0.23 s(-1), respectively at 300 K.