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Tuning the size and properties of ClyA nanopores assisted by directed evolution

Soskine, M., Biesemans, A., De Maeyer, M. & Maglia, G., 11-Sep-2013, In : Journal of the American Chemical Society. 135, 36, p. 13456-13463 8 p.

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  • Tuning the size and properties of ClyA nanopores assisted by directed evolution

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DOI

Nanopores have recently emerged as powerful tools in single-molecule investigations. Biological nanopores, however, have drawbacks, including a fixed size and limited stability in lipid bilayers. Inspired by the great success of directed evolution approaches in tailoring enzyme properties, in this work we evolved Cytolysin A from Salmonella typhi (ClyA) to a high level of soluble expression and desired electrical properties in lipid bilayers. Evolved ClyA nanopores remained open up to -150 mV applied potential, which allowed the detailed characterization of folded proteins by ionic current recordings. Remarkably, we also found that ClyA forms several nanopore species; among which we could isolate and characterize three nanopore types most likely corresponding to the 12mer, 13mer, and 14mer oligomeric forms of ClyA. Protein current blockades to the three ClyA nanopores showed that subnanometer variations in the diameter of nanopores greatly affect the recognition of analyte proteins.
Original languageEnglish
Pages (from-to)13456-13463
Number of pages8
JournalJournal of the American Chemical Society
Volume135
Issue number36
Publication statusPublished - 11-Sep-2013

    Keywords

  • solid-state nanopore, protein translocation, DNA translocation, E. coli, stranded DNA

ID: 36010469