Department of Molecular Biosciences, University of Texas at Austin, Austin, Texas, USA.
Center for Systems and Synthetic Biology, University of Texas at Austin, Austin, Texas, USA.
Protein Sci. 2021 May;30(5):1006-1021. doi: 10.1002/pro.4067. Epub 2021 Apr 9.
Electron microscopy (EM) continues to provide near-atomic resolution structures for well-behaved proteins and protein complexes. Unfortunately, structures of some complexes are limited to low- to medium-resolution due to biochemical or conformational heterogeneity. Thus, the application of unbiased systematic methods for fitting individual structures into EM maps is important. A method that employs co-evolutionary information obtained solely from sequence data could prove invaluable for quick, confident localization of subunits within these structures. Here, we incorporate the co-evolution of intermolecular amino acids as a new type of distance restraint in the integrative modeling platform in order to build three-dimensional models of atomic structures into EM maps ranging from 10-14 Å in resolution. We validate this method using four complexes of known structure, where we highlight the conservation of intermolecular couplings despite dynamic conformational changes using the BAM complex. Finally, we use this method to assemble the subunits of the bacterial holo-translocon into a model that agrees with previous biochemical data. The use of evolutionary couplings in integrative modeling improves systematic, unbiased fitting of atomic models into medium- to low-resolution EM maps, providing additional information to integrative models lacking in spatial data.
电子显微镜（EM）继续为行为良好的蛋白质和蛋白质复合物提供近原子分辨率的结构。不幸的是，由于生化或构象异质性，一些复合物的结构仅限于低分辨率到中分辨率。因此，应用无偏的系统方法将单个结构拟合到 EM 图谱中非常重要。一种仅从序列数据中获得共进化信息的方法，对于快速、自信地定位这些结构中的亚基可能非常有价值。在这里，我们将分子间氨基酸的共进化作为一种新的距离约束纳入集成建模平台中，以便将原子结构的三维模型构建到分辨率为 10-14Å 的 EM 图谱中。我们使用四个已知结构的复合物来验证这种方法，其中我们使用 BAM 复合物强调了尽管存在动态构象变化，但分子间耦合的保守性。最后，我们使用这种方法将细菌全转运通道的亚基组装成一个与以前生化数据一致的模型。在集成建模中使用进化耦合可以改善原子模型在中低分辨率 EM 图谱中的系统、无偏拟合，为缺乏空间数据的集成模型提供额外信息。
