Eliminating Critical Systematic Errors In Structural Biology With Next-Generation Simulation

项目总结/摘要 大分子晶体学中的数据收集易受重大系统误差的影响， 在许多系统上的成功数据收集，并最终限制了结果结构的准确性。创建 可以解释这些错误的仿真技术将在三个方面产生重大影响：1）解决 新的结构，更好地占辐射损伤，这是负责80%的失败异常 相位尝试，2）通过模拟非同构来改进多晶体平均，这将打开 通往任意增益的信号噪声，3）歧视激烈争议的替代解释，如 作为结合配体的存在或不存在，通过创建模拟与更现实的溶剂模型。到 为了从同步加速器获得“无损伤数据”，我们将首先校准辐射损伤曲线， 模型和DBP样本。利用这些曲线，我们将把辐射损伤的真实3D模型， 非长方体晶体（RADDOSE 3D）到我们的衍射图像模拟器（MLFSOM），以产生3D剂量 沿晶体沿着分布和照明贴图。这将导致新一代的波长- 晶体的相关吸收因子，以补充现有的吸收校正。在光束线处， 我们将使用锥形束在线X射线吸收射线照相术测量晶体的3D图， 的光束轮廓。这些进步将使我们能够在开放的环境中生成零剂量外推值。 格式，占实验晶体和光束的几何形状。为了改善多晶体平均，我们将 开始通过表征非同构如何作为湿度、辐射损伤和 功能状态通过更新经典的“克里克和马格努什”模拟的非同构与增加 复杂性，我们将开发一个奇异值分解方法参数非同构。使用 从这个分析得出的修正，我们将纠正非同构存在于多晶体 实验，能够确定新的结构，包括那些收集使用串行 下一代光源的晶体学。为了增强模拟，以便对 实验数据，我们将利用新的溶剂模型在大分子晶体学和小角度X- 射线散射我们的工作将创建标准协议，用于比较溶剂密度和替代品 解释，并定量评估每个模拟情况与真实的相比的可能性 大分子晶体学或SAXS数据。除了区分不同的解释， 实验数据，改善溶剂模型将提高理解如何大分子影响 并与其表面附近的其他分子相互作用。总的来说，我们希望消除这些障碍 关键的系统错误对方法发展和功能研究都是变革性的。

PROJECT SUMMARY/ABSTRACT Data collection in macromolecular crystallography is subject to significant systematic errors that prevent successful data collection on many systems and, ultimately, limit the accuracy of resulting structures. Creating simulation technologies that can account for these errors will have significant impact on three fronts: 1) solving new structures by better accounting for radiation damage, which is responsible for 80% of failed anomalous phasing attempts, 2) improving multi-crystal averaging by simulating non-isomorphism, which will open the gateway to arbitrary gains in signal-to-noise, 3) discriminating hotly contested alternative interpretations such as the presence or absence of a bound ligand, by creating simulations with more realistic solvent models. To move towards “damage-free data” from a synchrotron, we will start by calibrating radiation damage curves on model and DBP samples. Using these curves we will incorporate realistic 3D models of radiation damage to non-cuboid crystals (RADDOSE 3D) into our diffraction image simulator (MLFSOM) to yield a 3D Dose Distribution and Illumination map along the crystal. This will result in a new generation of wavelength- dependent absorption factors for the crystal to complement existing absorption corrections. At the beamline, we will measure a 3D map of the crystal using cone beam online x-ray absorption radiography and a 2D map of the beam profile. These advances will allow us to generate zero-dose extrapolation values, in an open format, that account for experimental crystal and beam geometry. To improve multi-crystal averaging, we will begin by characterizing how non-isomorphism varies as a function of humidity, radiation damage, and functional state. By updating the classic “Crick and Magdoff” simulations of non-isomorphism with increasing complexity, we will develop a singular value decomposition approach to parameterize non-isomorphism. Using the corrections derived from this analysis, we will correct the non-isomorphism present in multi-crystal experiments, enabling the determination of novel structures, including those collected using serial crystallography at next-generation light sources. To enable enhanced simulation for robust interpretation of experimental data, we will leverage new solvent models in macromolecular crystallography and small angle X- ray scattering. Our work will create standard protocols for comparing solvent density to alternative interpretations and to quantitatively assess how likely each simulated situation is compared to the real macromolecular crystallography or SAXS data. In addition to distinguishing between different interpretations of the experimental data, improving solvent models will enhance understanding of how macromolecules influence and interact with other molecules near their surface. Collectively, we expect the benefits of eliminating these critical systematic errors be transformative to both methods development and functional studies.