An essential intermediate in the folding of dihydrofolate reductase
An essential intermediate in the folding of dihydrofolate reductase
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DOI：
10.1073/pnas.100547697
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发表时间：
2000-05-23
影响因子：
11.1
通讯作者：
Jennings, PA
中科院分区：
文献类型：
--
作者：
Heidary, DK;O'Neill, JC;Jennings, PA
在pH7.8和15 ℃条件下，用停流荧光和吸收光谱法研究了大肠杆菌二氢叶酸还原酶的折叠。高度荧光的中间体的形成发生与弛豫时间范围在142和343毫秒之间，而停流吸收光谱使用甲氨蝶呤结合试验显示在这些时间范围内的天然状态的一个明显的滞后期。吸收动力学的滞后和快速折叠事件的缺乏表明，该中间体集合的形成是折叠反应中的必要步骤。
The folding of Escherichia coli dihydrofolate reductase was examined at pH 7.8 and 15 degrees C by using stopped-flow fluorescence and absorbance spectroscopies. The formation of a highly fluorescent intermediate occurs with relaxation times ranging between 142 and 343 msec, whereas stopped-flow absorbance spectroscopy using methotrexate binding assays shows a distinct lag phase during these time frames for the native state. The lag in absorbance kinetics and the lack of fast-track folding events indicate that the formation of this ensemble of intermediates is an obligatory step in the folding reaction.