Genetically encoded tags for direct synthesis of EM-visible gold nanoparticles in cells
| Autor: | Yuhua Li, Zhaodi Jiang, Gaihong Cai, Pei Zhao, Xiangbing Qi, Ying-Ying Wang, Li-Lin Du, Xinbin Cai, Yanyong Hu, Sitong Liu, Xiao-Man Liu, Yaqi Tang, Wanzhong He, Ying Liu, She Chen, Xiaobin Sun, Ming Li, Yan Liu, Xiumei Jin |
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| Rok vydání: | 2019 |
| Předmět: |
biology
Cell-Free System Chemistry Endoplasmic reticulum Metal Nanoparticles Cell Biology biology.organism_classification Biochemistry Fusion protein Spindle pole body Cell-free system Cell biology Microscopy Electron Microscopy Fluorescence Colloidal gold Spectrometry Mass Matrix-Assisted Laser Desorption-Ionization Organelle Schizosaccharomyces Humans Gold Nuclear pore Molecular Biology Biotechnology HeLa Cells |
| Zdroj: | Nature methods. 17(9) |
| ISSN: | 1548-7105 |
| Popis: | Genetically encoded tags for single-molecule imaging in electron microscopy (EM) are long-awaited. Here, we report an approach for directly synthesizing EM-visible gold nanoparticles (AuNPs) on cysteine-rich tags for single-molecule visualization in cells. We first uncovered an auto-nucleation suppression mechanism that allows specific synthesis of AuNPs on isolated tags. Next, we exploited this mechanism to develop approaches for single-molecule detection of proteins in prokaryotic cells and achieved an unprecedented labeling efficiency. We then expanded it to more complicated eukaryotic cells and successfully detected the proteins targeted to various organelles, including the membranes of endoplasmic reticulum (ER) and nuclear envelope, ER lumen, nuclear pores, spindle pole bodies and mitochondrial matrices. We further implemented cysteine-rich tag-antibody fusion proteins as new immuno-EM probes. Thus, our approaches should allow biologists to address a wide range of biological questions at the single-molecule level in cellular ultrastructural contexts. |
| Databáze: | OpenAIRE |
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