生物大分子,生命的基本组成部分,在自然环境中表现出动态结构。传统的结构确定技术通常通过仅捕获整体平均和时间平均的分子结构来过度简化这些复杂的构象光谱。解决这个差距,在这项工作中,我们将单目标散射采样(SOSS)方法的应用扩展到多种生物分子,包括RNA和蛋白质。我们的方法,称为“Bio-SOSS”,利用超短X射线脉冲捕获瞬时结构。在Bio-SOSS中,我们使用两个金纳米粒子(AuNPs)作为标签,在X射线散射信号中提供强烈的对比,以确保标记站点之间的精确距离确定。我们为RNA生成了假设的Bio-SOSS图像,蛋白质,和一个RNA-蛋白质复合物,每个在指定位置标记有两个AuNP。随后,为了验证Bio-SOSS的准确性,我们从图像中提取这些纳米颗粒标记之间的距离,并将它们与用于生成Bio-SOSS图像的实际值进行比较。具体来说,对于代表性RNA(1KXK),发现分子动力学快照和Bio-SOSS检索之间的距离差异的标准偏差最佳约为0.2µ,在最先进的X射线自由电子激光设施的实际实验条件下,通常在1µ内。此外,我们对各种实验因素,例如AuNP大小,X射线特性,和探测器的几何形状,影响Bio-SOSS的准确性。这项全面的研究突出了Bio-SOSS在准确捕获生物大分子的不同构象谱方面的实用性和潜力,为更深入地了解它们的动态性质铺平了道路。
Biological macromolecules, the fundamental building blocks of life, exhibit dynamic structures in their natural environment. Traditional structure determination techniques often oversimplify these multifarious conformational spectra by capturing only ensemble- and time-averaged molecular structures. Addressing this gap, in this work, we extend the application of the single-object scattering sampling (SOSS) method to diverse biological molecules, including RNAs and proteins. Our approach, referred to as \"Bio-SOSS\", leverages ultrashort X-ray pulses to capture instantaneous structures. In Bio-SOSS, we employ two gold nanoparticles (AuNPs) as labels, which provide strong contrast in the X-ray scattering signal, to ensure precise distance determinations between labeled sites. We generated hypothetical Bio-SOSS images for RNAs, proteins, and an RNA-protein complex, each labeled with two AuNPs at specified positions. Subsequently, to validate the accuracy of Bio-SOSS, we extracted distances between these nanoparticle labels from the images and compared them with the actual values used to generate the Bio-SOSS images. Specifically, for a representative RNA (1KXK), the standard deviation in distance discrepancies between molecular dynamics snapshots and Bio-SOSS retrievals was found to be optimally around 0.2 Å, typically within 1 Å under practical experimental conditions at state-of-the-art X-ray free-electron laser facilities. Furthermore, we conducted an in-depth analysis of how various experimental factors, such as AuNP size, X-ray properties, and detector geometry, influence the accuracy of Bio-SOSS. This comprehensive investigation highlights the practicality and potential of Bio-SOSS in accurately capturing the diverse conformation spectrum of biological macromolecules, paving the way for deeper insights into their dynamic natures.