Oxford Particle Imaging Centre, Division of Structural Biology, University of Oxford, Wellcome Centre for Human Genetics, Roosevelt Drive, Oxford UK OX3 7BN; Department of Physics, University of Oxford, Clarendon Laboratory, Parks Road, Oxford UK OX1 3PU; Centre for Structural Systems Biology, Heinrich-Pette-Institut, Leibniz-Institut für Experimentelle Virologie, Notkestrasse 85, 22607 Hamburg, Germany.
Oxford Particle Imaging Centre, Division of Structural Biology, University of Oxford, Wellcome Centre for Human Genetics, Roosevelt Drive, Oxford UK OX3 7BN; Centre for Structural Systems Biology, Heinrich-Pette-Institut, Leibniz-Institut für Experimentelle Virologie, Notkestrasse 85, 22607 Hamburg, Germany.
Cell. 2021 Feb 18;184(4):1110-1121.e16. doi: 10.1016/j.cell.2021.01.033.
Electron cryotomography (cryoET), an electron cryomicroscopy (cryoEM) modality, has changed our understanding of biological function by revealing the native molecular details of membranes, viruses, and cells. However, identification of individual molecules within tomograms from cryoET is challenging because of sample crowding and low signal-to-noise ratios. Here, we present a tagging strategy for cryoET that precisely identifies individual protein complexes in tomograms without relying on metal clusters. Our method makes use of DNA origami to produce "molecular signposts" that target molecules of interest, here via fluorescent fusion proteins, providing a platform generally applicable to biological surfaces. We demonstrate the specificity of signpost origami tags (SPOTs) in vitro as well as their suitability for cryoET of membrane vesicles, enveloped viruses, and the exterior of intact mammalian cells.
电子低温断层摄影术（cryoET），一种电子低温显微镜（cryoEM）模式，通过揭示膜、病毒和细胞的天然分子细节，改变了我们对生物功能的理解。然而，由于样品拥挤和低信噪比，从 cryoET 的断层图像中识别单个分子具有挑战性。在这里，我们提出了一种 cryoET 的标记策略，该策略无需依赖金属簇即可精确识别断层图像中的单个蛋白质复合物。我们的方法利用 DNA 折纸术产生“分子路标”，靶向感兴趣的分子，这里通过荧光融合蛋白，为生物表面提供了一个普遍适用的平台。我们证明了路标折纸标签（SPOTs）在体外的特异性以及它们在膜泡、包膜病毒和完整哺乳动物细胞外的 cryoET 中的适用性。
Zotero 插件
