{
  "abstract": "Positive reversibly switchable fluorescent proteins (+rsFPs) are genetically encoded fluorescent labels allowing super-resolution microscopy without the fading inherent in the better established negative (−) rsFPs. Typically, switching is initiated by an ultrafast excited-state isomerization reaction. Here, we investigate the ultrafast photophysics of four structurally similar variants of the archetypal +rsFP Padron, including Padron0.9, Padron2, Kohinoor, and a newly reported “reverse” Kohinoor mutant (Kohinoor S176F) using ultrafast optical and vibrational spectroscopy. Padron0.9 exhibits an efficient excited-state proton transfer (ESPT) reaction in parallel with photoisomerization, whereas Kohinoor and Padron2 do not. Remarkably, introducing the single reverse mutation S176F in Kohinoor restores Padron-like ESPT dynamics, even though the mutation is remote from the proton donor. This result demonstrates that a single amino acid residue near to, but not in direct H-bonded interaction with, the chromophore can govern the fate of excited states. TRIR measurements reveal distinct changes in the local hydrogen-bonding network and chromophore–protein interactions associated with ESPT-active variants and further reveal a key role for H-bonding between the chromophore and an arginine residue through a study of the R66K mutant of Padron0.9. Overall, these results show that subtle mutations shift the excited state between proton transfer and photoisomerization pathways, providing mechanistic insight into photoswitching in rsFPs, thus guiding the design of improved fluorescent proteins for superresolution bioimaging.",
  "abstract_shown": true,
  "affiliations": [
    "School of Chemistry, University of East Anglia, Norwich NR4 7TJ, U.K.",
    "Department of Chemistry, Stony Brook University, Stony Brook, New York 11794, United States",
    "Central Laser Facility, Research Complex at Harwell, Rutherford Appleton Laboratory, Didcot OX11 0QX, U.K.",
    "Department of Biophysics, Medical School, University of Pecs, 7624 Pecs, Hungary"
  ],
  "article_type": "research article",
  "arxiv": null,
  "author_total": 9,
  "authors": [
    {
      "affs": [
        0
      ],
      "name": "Anam Fatima",
      "orcid": "0000-0003-2388-4390"
    },
    {
      "affs": [
        1
      ],
      "name": "Danielle Rosenberger"
    },
    {
      "affs": [
        1
      ],
      "name": "YongLe He",
      "orcid": "0000-0001-7274-535X"
    },
    {
      "affs": [
        1
      ],
      "name": "James N. Iuliano",
      "orcid": "0000-0003-1213-3292"
    },
    {
      "affs": [
        2
      ],
      "name": "Gregory M. Greetham",
      "orcid": "0000-0002-1852-3403"
    },
    {
      "affs": [
        2
      ],
      "name": "Partha Malakar",
      "orcid": "0000-0001-6874-7010"
    },
    {
      "affs": [
        3
      ],
      "name": "Andras Lukacs",
      "orcid": "0000-0001-8841-9823"
    },
    {
      "affs": [
        1
      ],
      "name": "Peter J. Tonge",
      "orcid": "0000-0003-1606-3471"
    },
    {
      "affs": [
        0
      ],
      "name": "Stephen R. Meech",
      "orcid": "0000-0001-5561-2782"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/653phdpq3gnez.bib",
  "cited_by": 0,
  "doi": "10.1021/acs.jpclett.6c02808",
  "fulltext_url": "https://pmc.ncbi.nlm.nih.gov/articles/PMC13726473/",
  "id": "653phdpq3gnez",
  "issns": [
    "1948-7185"
  ],
  "landing_url": "https://doi.org/10.1021/acs.jpclett.6c02808",
  "language": "en",
  "licence": "CC-BY-4.0",
  "licence_label": "CC BY 4.0",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "heading",
      "Letter"
    ],
    [
      "heading",
      "Physical Insights into Light Interacting with Matter"
    ],
    [
      "openalex",
      "1304|Advanced Fluorescence Microscopy Techniques"
    ]
  ],
  "openalex": "W7223035530",
  "pdf_url": null,
  "pmcid": "PMC13726473",
  "pmid": "42954849",
  "published": "2026-10-20",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/653phdpq3gnez.ris",
  "sources": [
    {
      "first_seen": "2026-11-05T12:22:49Z",
      "native_id": "PMC13726473",
      "source": "pmc",
      "url": "https://doi.org/10.1021/acs.jpclett.6c02808"
    }
  ],
  "subjects": [
    {
      "field": "biochemistry",
      "origin": "openalex:1304|Advanced Fluorescence Microscopy Techniques",
      "subfield": "biophysics"
    }
  ],
  "title": "Remote Control of Excited-State Chemistry in Positive Reversibly Switchable Fluorescent Proteins",
  "title_text": "Remote Control of Excited-State Chemistry in Positive Reversibly Switchable Fluorescent Proteins",
  "type": "peer-reviewed",
  "updated_at": "2026-11-06T12:22:26Z",
  "url": "https://research.draconitic.com/paper/653phdpq3gnez",
  "venue": "The Journal of Physical Chemistry Letters",
  "versions": []
}