Rapid discovery and evolution of nanosensors containing fluorogenic amino acids
Binding-activated optical sensors are powerful tools for imaging, diagnostics, and biomolecular sensing. However, biosensor discovery is slow and requires tedious steps in rational design, screening, and characterization. Here we report on a platform that streamlines biosensor discovery and unlocks directed nanosensor evolution through genetically encodable fluorogenic amino acids (FgAAs). Building on the classical knowledge-based semisynthetic approach, we engineer ~15 kDa nanosensors that recognize specific proteins, peptides, and small molecules with up to 100-fold fluorescence increases and subsecond kinetics, allowing real-time and wash-free target sensing and live-cell bioimaging. An optimized genetic code expansion chemistry with FgAAs further enables rapid (~3 h) ribosomal nanosensor discovery via the cell-free translation of hundreds of candidates in parallel and directed nanosensor evolution with improved variant-specific sensitivities (up to ~250-fold) for SARS-CoV-2 antigens. Altogether, this platform could accelerate the discovery of fluorogenic nanosensors and pave the way to modify proteins with other non-standard functionalities for diverse applications.
Authors
- Jason D. Galpin (ORCID: https://orcid.org/0000-0003-1923-5312)
- William Bedell
- Thomas M. Bartlett (ORCID: https://orcid.org/0000-0001-9730-3638)
- Fabio De Moliner
- Isaac Han (ORCID: https://orcid.org/0000-0003-4785-9042)
- Marc Vendrell (ORCID: https://orcid.org/0000-0002-5392-9740)
- Helena de Puig
- Subhrajit Rout
- Abigail E Reese (ORCID: https://orcid.org/0009-0003-6915-0402)
- Lindsey R. Robinson‐McCarthy (ORCID: https://orcid.org/0000-0002-8066-2770)
- Christopher A. Ahern (ORCID: https://orcid.org/0000-0002-7975-2744)
- Jonathan Rittichier (ORCID: https://orcid.org/0000-0002-4264-8628)
- Allison Flores (ORCID: https://orcid.org/0000-0002-2655-8323)
- Jorge A. Marchand (ORCID: https://orcid.org/0000-0002-7765-610X)
- James J. Collins (ORCID: https://orcid.org/0000-0002-5560-8246)
- David Z. Rudner (ORCID: https://orcid.org/0000-0002-0236-7143)
- Thomas G. Bernhardt (ORCID: https://orcid.org/0000-0003-3566-7756)
- Erkin Kuru (ORCID: https://orcid.org/0000-0002-2356-0858)
- George M. Church
- Sylvie G. Bernier
Institutions
- Broad Institute (US)
- University of Iowa (US)
- New York State Department of Health (US)
- Wadsworth Center (US)
- Howard Hughes Medical Institute (US)
- Harvard University (US)
- University of Washington (US)
- Dana-Farber/Harvard Cancer Center (US)
- Pediatrics and Genetics (US)
- Centre for Inflammation Research (GB)
- Center for Systems Biology (US)
- Massachusetts Institute of Technology (US)
Publication Details
- Journal
- Nature Communications
- Published
- 2024-09-05
- DOI
- https://doi.org/10.1038/s41467-024-50956-z
- Citations
- 7
- Primary Topic
- Advanced biosensing and bioanalysis techniques
- Type
- article
- Field-Weighted Citation Impact
- 1.64
Funders
- U.S. Department of Energy
- Life Sciences Research Foundation
- Hansjörg Wyss Institute for Biologically Inspired Engineering, Harvard University