Ratner V, Sinev M, Haas E
Department of Life Sciences, Bar Ilan University, Ramat-Gan, 52900, Israel.
J Mol Biol. 2000 Jun 23;299(5):1363-71. doi: 10.1006/jmbi.2000.3814.
A method for determination of transient (on the millisecond timescale) intramolecular distance distributions (IDDs) by time-resolved dynamic non-radiative excitation energy transfer measurements was developed. The time-course of the development of the IDD between residues 73 and 203 in the CORE domain of Escherichia coli adenylate kinase throughout refolding from the GuHCl-induced denatured state was determined. The mean of the apparent IDD reduced to a value close to its magnitude in the native protein, within 2 ms (the dead-time of the instrument). At that time the width of that distribution was rather large (16+/-2 A). The large width implies that the intramolecular diffusion coefficient of the labeled segment does not exceed 10(-7) cm(2)/second. In a second slower phase of the refolding transition, the width was reduced to its native value (6+/-4 A).
开发了一种通过时间分辨动态非辐射激发能量转移测量来测定瞬态（毫秒时间尺度）分子内距离分布（IDDs）的方法。确定了大肠杆菌腺苷酸激酶核心结构域中73位和203位残基之间的IDD在从盐酸胍诱导的变性状态重新折叠过程中的发展时间进程。表观IDD的平均值在2毫秒内（仪器的死时间）降至接近其在天然蛋白质中的大小的值。此时，该分布的宽度相当大（16±2 Å）。大宽度意味着标记片段的分子内扩散系数不超过10^(-7) cm²/秒。在重新折叠转变的第二个较慢阶段，宽度减小到其天然值（6±4 Å）。
