Increasingly, cryo-electron microscopy (cryo-EM) is used to determine the structures of RNA–protein assemblies, but nearly all maps determined with this method have biologically important regions where the local resolution does not permit RNA coordinate tracing. To address these omissions, we present de novo ribonucleoprotein modeling in real space through assembly of fragments together with experimental density in Rosetta (DRRAFTER). We show that DRRAFTER recovers near-native models for a diverse benchmark set of RNA–protein complexes including the spliceosome, mitochondrial ribosome, and CRISPR–Cas9–sgRNA complexes; rigorous blind tests include yeast U1 snRNP and spliceosomal P complex maps. Additionally, to aid in model interpretation, we present a method for reliable in situ estimation of DRRAFTER model accuracy. Finally, we apply DRRAFTER to recently determined maps of telomerase, the HIV-1 reverse transcriptase initiation complex, and the packaged MS2 genome, demonstrating the acceleration of accurate model building in challenging cases.

冷冻电子显微镜 (cryo-EM) 越来越多地用于确定 RNA-蛋白质组装体的结构，但几乎所有用这种方法确定的图谱都具有生物学上重要的区域，这些区域的局部分辨率不允许 RNA 坐标追踪。为了解决这些遗漏，我们通过在 Rosetta (DRRAFTER) 中组装片段和实验密度，在真实空间中进行从头核糖核蛋白建模。我们表明，DRRAFTER 恢复了多种 RNA-蛋白质复合物基准组的近乎天然模型，包括剪接体、线粒体核糖体和 CRISPR-Cas9-sgRNA 复合物；严格的盲测包括酵母 U1 snRNP 和剪接体 P 复合物图谱。此外，为了帮助模型解释，我们提出了一种可靠的原位估计 DRRAFTER 模型精度的方法。最后，我们将 DRRAFTER 应用于最近确定的端粒酶、HIV-1 逆转录酶起始复合物和包装的 MS2 基因组图谱，证明了在具有挑战性的情况下加速了准确模型的构建。