Title
Full-length single-molecule protein fingerprinting
Author
Filius, M. (TU Delft BN/Chirlmin Joo Lab; Kavli institute of nanoscience Delft) 
van Wee, R.G. (TU Delft BN/Chirlmin Joo Lab; Kavli institute of nanoscience Delft)
de Lannoy, C.V. (TU Delft BN/Chirlmin Joo Lab; Wageningen University & Research; Kavli institute of nanoscience Delft)
Westerlaken, I. (TU Delft BT/Industriele Microbiologie; Kavli institute of nanoscience Delft)
Li, Zeshi (Kavli institute of nanoscience Delft)
Kim, S.H. (TU Delft BN/Chirlmin Joo Lab; Ewha Womans University; Kavli institute of nanoscience Delft)
de Agrela Pinto, C. (TU Delft BN/Arjen Jakobi Lab; Kavli institute of nanoscience Delft)
Wu, Yunfei (Universiteit Utrecht)
Boons, Geert-Jan (Universiteit Utrecht)
Pabst, Martin (TU Delft BT/Environmental Biotechnology)
de Ridder, Dick (Wageningen University & Research)
Joo, C. (TU Delft BN/Chirlmin Joo Lab; Kavli institute of nanoscience Delft)
Date
2024
Abstract
Proteins are the primary functional actors of the cell. While proteoform diversity is known to be highly biologically relevant, current protein analysis methods are of limited use for distinguishing proteoforms. Mass spectrometric methods, in particular, often provide only ambiguous information on post-translational modification sites, and sequences of co-existing modifications may not be resolved. Here we demonstrate fluorescence resonance energy transfer (FRET)-based single-molecule protein fingerprinting to map the location of individual amino acids and post-translational modifications within single full-length protein molecules. Our data show that both intrinsically disordered proteins and folded globular proteins can be fingerprinted with a subnanometer resolution, achieved by probing the amino acids one by one using single-molecule FRET via DNA exchange. This capability was demonstrated through the analysis of alpha-synuclein, an intrinsically disordered protein, by accurately quantifying isoforms in mixtures using a machine learning classifier, and by determining the locations of two O-GlcNAc moieties. Furthermore, we demonstrate fingerprinting of the globular proteins Bcl-2-like protein 1, procalcitonin and S100A9. We anticipate that our ability to perform proteoform identification with the ultimate sensitivity may unlock exciting new venues in proteomics research and biomarker-based diagnosis
To reference this document use:
http://resolver.tudelft.nl/uuid:46f7dd57-fc54-415c-8793-a6696014963f
DOI
https://doi.org/10.1038/s41565-023-01598-7
Embargo date
2024-08-13
ISSN
1748-3395
Source
Nature Nanotechnology
Bibliographical note
Green Open Access added to TU Delft Institutional Repository 'You share, we take care!' - Taverne project https://www.openaccess.nl/en/you-share-we-take-care Otherwise as indicated in the copyright section: the publisher is the copyright holder of this work and the author uses the Dutch legislation to make this work public.
Part of collection
Institutional Repository
Document type
journal article
Rights
© 2024 M. Filius, R.G. van Wee, C.V. de Lannoy, I. Westerlaken, Zeshi Li, S.H. Kim, C. de Agrela Pinto, Yunfei Wu, Geert-Jan Boons, Martin Pabst, Dick de Ridder, C. Joo