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Electroporated recombinant proteins as tools for in vivo functional complementation, imaging and chemical biology
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2019 (English)In: eLIFE, E-ISSN 2050-084X, Vol. 8, article id e48287Article in journal (Refereed) Published
Abstract [en]

Delivery of native or chemically modified recombinant proteins into mammalian cells shows promise for functional investigations and various technological applications, but concerns that sub-cellular localization and functional integrity of delivered proteins may be affected remain high. Here, we surveyed batch electroporation as a delivery tool for single polypeptides and multi-subunit protein assemblies of the kinetochore, a spatially confined and well-studied subcellular structure. After electroporation into human cells, recombinant fluorescent Ndc80 and Mis12 multi-subunit complexes exhibited native localization, physically interacted with endogenous binding partners, and functionally complemented depleted endogenous counterparts to promote mitotic checkpoint signaling and chromosome segregation. Farnesylation is required for kinetochore localization of the Dynein adaptor Spindly. In cells with chronically inhibited farnesyl transferase activity, in vitro farnesylation and electroporation of recombinant Spindly faithfully resulted in robust kinetochore localization. Our data show that electroporation is well-suited to deliver synthetic and chemically modified versions of functional proteins, and, therefore, constitutes a promising tool for applications in chemical and synthetic biology.

Place, publisher, year, edition, pages
ELIFE SCIENCES PUBLICATIONS LTD , 2019. Vol. 8, article id e48287
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
URN: urn:nbn:se:umu:diva-161999DOI: 10.7554/eLife.48287ISI: 000477003900001PubMedID: 31310234OAI: oai:DiVA.org:umu-161999DiVA, id: diva2:1342235
Available from: 2019-08-13 Created: 2019-08-13 Last updated: 2019-08-13Bibliographically approved

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Wu, Yao-Wen

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Alex, AmalPiano, ValentinaStuiver, MarchelWu, Yao-Wen
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Department of Chemistry
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Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)

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CiteExportLink to record
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Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
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  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
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  • Other locale
More languages
Output format
  • html
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