Bioactivated and PEG-Protected Circa 2 nm Gold Nanoparticles for in Cell Labelling and Cryo-Electron Microscopy


Groysbeck, Nadja, Hanss, Victor, Donzeau, Mariel, Strub, Jean-Marc, Cianferani, Sarah, Spehner, Daniele, Bahri, Mounib ORCID: 0000-0002-8336-9158, Ersen, Ovidiu, Eltsov, Mikhael, Schultz, Patrick et al (show 1 more authors) (2023) Bioactivated and PEG-Protected Circa 2 nm Gold Nanoparticles for in Cell Labelling and Cryo-Electron Microscopy. SMALL METHODS, 7 (6). e2300098-.

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Abstract

Advances in cryo-electron microscopy (EM) enable imaging of protein assemblies within mammalian cells in a near native state when samples are preserved by cryogenic vitrification. To accompany this progress, specialized EM labelling protocols must be developed. Gold nanoparticles (AuNPs) of 2 nm are synthesized and functionalized to bind selected intracellular targets inside living human cells and to be detected in vitreous sections. As a proof of concept, thioaminobenzoate-, thionitrobenzoate-coordinated gold nanoparticles are functionalized on their surface with SV40 Nuclear Localization Signal (NLS)-containing peptides and 2 kDa polyethyleneglycols (PEG) by thiolate exchange to target the importin-mediated nuclear machinery and facilitate cytosolic diffusion by shielding the AuNP surface from non-specific binding to cell components, respectively. After delivery by electroporation into the cytoplasm of living human cells, the PEG-coated AuNPs diffuse freely in the cytoplasm but do not enter the nucleus. Incorporation of NLS within the PEG coverage promotes a quick nuclear import of the nanoparticles in relation to the density of NLS onto the AuNPs. Cryo-EM of vitreous cell sections demonstrate the presence of 2 nm AuNPs as single entities in the nucleus. Biofunctionalized AuNPs combined with live-cell electroporation procedures are thus potent labeling tools for the identification of macromolecules in cellular cryo-EM.

Item Type: Article
Uncontrolled Keywords: bioconjugation, electroporation, gold labeling, gold particles, nuclear import, transduction
Divisions: Faculty of Science and Engineering > School of Engineering
Depositing User: Symplectic Admin
Date Deposited: 14 May 2023 19:16
Last Modified: 19 Jul 2023 19:24
DOI: 10.1002/smtd.202300098
Open Access URL: https://doi.org/10.1002/smtd.202300098
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URI: https://livrepository.liverpool.ac.uk/id/eprint/3170346
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