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An engineered CD81-based combinatorial library for selecting recombinant binders to cell surface proteins: Laminin binding CD81 enhances cellular uptake of extracellular vesicles

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DOI

  • Stefan Vogt, Austrian Centre of Industrial Biotechnology GmbH, University of Natural Resources and Life Sciences, Vienna
  • ,
  • Madhusudhan Reddy Bobbili, University of Natural Resources and Life Sciences, Vienna, Ludwig Boltzmann Institute for Experimental and Clinical Traumatology
  • ,
  • Gerhard Stadlmayr, University of Natural Resources and Life Sciences, Vienna
  • ,
  • Katharina Stadlbauer, University of Natural Resources and Life Sciences, Vienna
  • ,
  • Jørgen Kjems
  • Florian Rüker, University of Natural Resources and Life Sciences, Vienna
  • ,
  • Johannes Grillari, University of Natural Resources and Life Sciences, Vienna, Ludwig Boltzmann Institute for Experimental and Clinical Traumatology
  • ,
  • Gordana Wozniak-Knopp, University of Natural Resources and Life Sciences, Vienna

The research of extracellular vesicles (EVs) has boomed in the last decade, with the promise of them functioning as target-directed drug delivery vehicles, able to modulate proliferation, migration, differentiation, and other properties of the recipient cell that are vital for health of the host organism. To enhance the ability of their targeted delivery, we employed an intrinsically overrepresented protein, CD81, to serve for recognition of the desired target antigen. Yeast libraries displaying mutant variants of the large extracellular loop of CD81 have been selected for binders to human placental laminin as an example target. Their specific interaction with laminin was confirmed in a mammalian display system. Derived sequences were reformatted to full-length CD81 and expressed in EVs produced by HeLa cells. These EVs were examined for the presence of the recombinant protein and were shown to exhibit an enhanced uptake into laminin-secreting mammalian cell lines. For the best candidate, the specificity of antigen interaction was demonstrated with a competition experiment. To our knowledge, this is the first example of harnessing an EV membrane protein as mediator of de novo target antigen recognition via in vitro molecular evolution, opening horizons to a broad range of applications in various therapeutic settings.

Original languageEnglish
Article numbere12139
JournalJournal of Extracellular Vesicles
Volume10
Issue11
ISSN2001-3078
DOIs
Publication statusPublished - Sep 2021

    Research areas

  • antigen-binding CD81, directed evolution, extracellular vesicle uptake, targeted extracellular vesicles, yeast display

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