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Extension-Dependent Drift Velocity and Diffusion (DrDiff) Directly Reconstructs the Folding Free Energy Landscape of Atomic Force Microscopy Experiments.
The Journal of Physical Chemistry Letters ( IF 5.7 ) Pub Date : 2020-01-17 , DOI: 10.1021/acs.jpclett.9b02146
Frederico Campos Freitas 1 , Ronaldo Junio de Oliveira 1
Affiliation  

Two equilibrium force microscopy trajectories [q(t)] of high-precision single-molecule spectroscopy assays were analyzed: the pulling of an HIV RNA hairpin and of a 3-aa sequence of the bacteriorhodopsin membrane protein. Both present hundreds of two-state folding transitions, and their free-energy [F(q)] landscapes were previously obtained by deconvolving time signals with the inverse Boltzmann and pfold methods. In this letter, the two F profiles were reconstructed directly from the measured time-series by the drift-diffusion (DrDiff) framework that characterized the effective conformational drift-velocity [v(q)] and diffusion [D(q)] coefficients. The two thermodynamic F profiles reconstructed with DrDiff directly from q(t) were in good agreement with those previously obtained from the deconvolved time signals. q(t) trajectories simulated with a two-dimensional framework in which the diffusion coefficient of the pulling setup (q coordinate) differed from the molecule (x coordinate) were also analyzed by DrDiff. The performance in reconstructing F was investigated in different conditions of diffusion anisotropy in the simulated time-series using Brownian dynamics. In addition, recently developed theories were used in order to evaluate the quality of the analysis performed in the experimental time series: the memory effects and the intrinsic biomolecular dynamic properties after connecting the probe to the molecule. With the 2-dimensional diffusive models and the additional analyses, it is proposed that the different physical regimes imposed by the stiffer probes of the two biomolecules will have an impact in the measured extension-dependent D and, thus, in the reconstruction of F by DrDiff. Stiffer AFM probes may reflect the molecular behavior more faithfully and reconstruction of F might be more successful. The reported quantities extracted directly from q(t) highlights the current state of the biomolecule characterization by force spectroscopy experiments: it is still challenging despite the recent advances, yet it is very promising.

中文翻译:

扩展相关的漂移速度和扩散(DrDiff)直接重建了原子力显微镜实验的折叠自由能态。

高精度单分子光谱分析的两个平衡力显微镜轨迹[q(t)]进行了分析:HIV RNA发夹的拉动和细菌视紫红质膜蛋白的3-aa序列的拉动。两者都存在数百个二态折叠跃迁,并且它们的自由能[F(q)]先前是通过使用逆玻耳兹曼和pfold方法对时间信号进行反卷积获得的。在这封信中,两个F轮廓是通过漂移扩散(DrDiff)框架直接从测量的时间序列重建的,该框架描述了有效构象漂移速度[v(q)]和扩散[D(q)]系数。直接从q(t)用DrDiff重建的两个热力学F曲线与先前从反卷积时间信号中获得的曲线吻合得很好。还通过DrDiff分析了二维框架中模拟的q(t)轨迹,其中牵引设置的扩散系数(q坐标)与分子的扩散系数(x坐标)不同。在模拟的时间序列中,利用布朗动力学研究了在扩散各向异性不同条件下重建F的性能。另外,为了评估在实验时间序列中进行的分析的质量,使用了最近开发的理论:将探针连接到分子后的记忆效应和固有的生物分子动力学特性。利用二维扩散模型和附加分析,建议由两种生物分子的较硬探针施加的不同物理方案将对测得的延伸依赖性D产生影响,因此,在DrDiff重建F中。较硬的AFM探针可能更忠实地反映分子行为,并且F的重建可能更成功。直接从q(t)中提取的报告数量突出了通过力能谱实验表征生物分子的当前状态:尽管最近取得了一些进展,但仍具有挑战性,但非常有希望。
更新日期:2020-01-17
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