Resolving Chemical Modifications to a Single Amino Acid within a Peptide Using a Biological Nanopore

Laura Restrepo-Pérez, Gang Huang, Peggy R. Bohländer, Nathalie Worp, Rienk Eelkema, Giovanni Maglia, Chirlmin Joo, Cees Dekker

Research output: Contribution to journalArticlepeer-review

66 Scopus citations


While DNA sequencing is now amply available, fast, and inexpensive, protein sequencing remains a tremendous challenge. Nanopores may allow for developing a protein sequencer with single-molecule capabilities. As identification of 20 different amino acids currently presents an unsurmountable challenge, fingerprinting schemes are pursued, in which only a subset of amino acids is labeled and detected. This requires modification of amino acids with chemical structures that generate a distinct nanopore ionic current signal. Here, we use a model peptide and the fragaceatoxin C nanopore to characterize six potential tags for a fingerprinting approach using nanopores. We find that labeled and unlabeled proteins can be clearly distinguished and that sensitive detection is obtained for labels with a spectrum of different physicochemical properties such as mass (427-1275 Da), geometry, charge, and hydrophobicity. Additionally, information about the position of the label along the peptide chain can be obtained from individual current-blockade event features. The results represent an important advance toward the development of a single-molecule protein-fingerprinting device with nanopores.

Original languageEnglish
Pages (from-to)13668-13676
Number of pages9
JournalACS Nano
Issue number12
StatePublished - 24 Dec 2019

Bibliographical note

Funding Information:
We would like to thank Niels van den Broek, Robert Cordfunke, Mike Filius, Marek Noga, Sonja Schmid, and Chun Heung Wong for fruitful discussions. This work was supported by the European Research Council Advanced Grant SynDiv (No. 669598) to C.D., by The Netherlands Organisation for Scientific Research (NWO/OCW) through the NanoFront and BaSyC grants, and by the Foundation for Fundamental Research on Matter (Vrije Programma SMPS) to C.J., C.D., R.E., P.R.B., and G.M.

Publisher Copyright:
© 2019 American Chemical Society.


  • amino acid labeling
  • biological nanopores
  • nanopore
  • protein analysis
  • protein fingerprinting
  • single-molecule protein sequencing


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