{
  "abstract": "Herein, we present three crystal structures of the most complex knotted protein to date, an 83 knot with eight crossings, which belongs to a clasp knot family. The fold of this protein is also unique, and we call it the EIGHT fold (Extraordinarily Intricate Globular Helical Tangled fold). The 83 knot is found only in a certain group of bacteria, where it is strictly conserved, despite sequence identities as low as 10%. The 83 structure shows a high level of internal symmetry, with structurally analogous halves. It is possible that the EIGHT fold evolved from a single unknotted “half” that duplicated and formed a complex protein. Based on an in vitro study of three selected 83 proteins, we estimated their thermal and thermodynamic stability, and showed that they follow a two-state unfolding model under equilibrium conditions and their refolding occurs to a compact, nonnative state. Unfolding kinetics of 83 knotted proteins reveal a complex, multiphase process spanning milliseconds to days. Six kinetic phases were resolved using stopped-flow fluorescence and far-ultraviolet absorbance circular dichroism spectroscopy, highlighting both rapid early events and slower rearrangements in secondary and tertiary structures. Finally, with a structure-based model, we showed that proteins with an 83 knot can self-tie through a 61 knotted intermediate state. They can also self-untie, albeit via an additional intermediate knot such as 41. Nevertheless, these proteins do not seem to fit with current folding theories known to date and require the development of a new approach.",
  "abstract_shown": true,
  "affiliations": [
    "Centre of New Technologies, University of Warsaw, Banacha 2C St., Warszawa 02-097, Poland",
    "Jerzy Haber Institute of Catalysis and Surface Chemistry, Polish Academy of Sciences, Krakow 30-239, Poland",
    "Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Warsaw 02-089, Poland",
    "Faculty of Chemistry, University of Warsaw, Warsaw 02-093, Poland",
    "Yusuf Hamied Department of Chemistry, University of Cambridge, Cambridge, CB2 1EW, United Kingdom"
  ],
  "article_type": "research article",
  "arxiv": null,
  "author_total": 12,
  "authors": [
    {
      "affs": [
        0
      ],
      "name": "Mariusz Mozajew",
      "orcid": "0000-0001-6157-2860"
    },
    {
      "affs": [
        0
      ],
      "name": "Maciej Sikora",
      "orcid": "0009-0003-0289-276X"
    },
    {
      "affs": [
        0
      ],
      "name": "Mateusz Fortunka",
      "orcid": "0000-0002-1756-2734"
    },
    {
      "affs": [
        0
      ],
      "name": "Agata P. Perlinska"
    },
    {
      "affs": [
        0,
        1
      ],
      "name": "Anna Kluza"
    },
    {
      "affs": [
        0
      ],
      "name": "Szymon Niewieczerzal",
      "orcid": "0000-0002-7563-7212"
    },
    {
      "affs": [
        0
      ],
      "name": "Fernando Bruno da Silva",
      "orcid": "0000-0002-0285-8700"
    },
    {
      "affs": [
        2
      ],
      "name": "Maciej Lukaszewicz"
    },
    {
      "affs": [
        0
      ],
      "name": "Julia A. Sikorska",
      "orcid": "0000-0002-4117-9225"
    },
    {
      "affs": [
        0,
        3
      ],
      "name": "Lena J. Mikulska",
      "orcid": "0009-0009-9052-0037"
    },
    {
      "affs": [
        4
      ],
      "name": "Sophie E. Jackson",
      "orcid": "0000-0002-7470-9800"
    },
    {
      "affs": [
        0
      ],
      "name": "Joanna I. Sulkowska",
      "orcid": "0000-0003-2452-0724"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/fj84gf4b9qtnv.bib",
  "cited_by": 0,
  "doi": "10.1073/pnas.2523017123",
  "fulltext_url": "https://pmc.ncbi.nlm.nih.gov/articles/PMC13715821/",
  "id": "fj84gf4b9qtnv",
  "issns": [
    "0027-8424",
    "1091-6490"
  ],
  "landing_url": "https://doi.org/10.1073/pnas.2523017123",
  "language": "en",
  "licence": "CC-BY-NC-ND-4.0",
  "licence_label": "CC BY-NC-ND 4.0",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by-nc-nd/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "heading",
      "408"
    ],
    [
      "heading",
      "Biological Sciences"
    ],
    [
      "heading",
      "Biophysics and Computational Biology"
    ],
    [
      "openalex",
      "1312|Protein Structure and Dynamics"
    ]
  ],
  "openalex": "W7222939779",
  "pdf_url": null,
  "pmcid": "PMC13715821",
  "pmid": "42896058",
  "published": "2026-10-19",
  "publisher": "National Academy of Sciences",
  "ris": "https://research.draconitic.com/paper/fj84gf4b9qtnv.ris",
  "sources": [
    {
      "first_seen": "2026-11-01T12:23:01Z",
      "native_id": "PMC13715821",
      "source": "pmc",
      "url": "https://doi.org/10.1073/pnas.2523017123"
    }
  ],
  "subjects": [
    {
      "field": "biochemistry",
      "origin": "openalex:1312|Protein Structure and Dynamics",
      "subfield": "molecular-biology"
    }
  ],
  "title": "Unique features of EIGHT fold—Proteins with the most complex 83 knotted topology",
  "title_text": "Unique features of EIGHT fold—Proteins with the most complex 83 knotted topology",
  "type": "peer-reviewed",
  "updated_at": "2026-11-01T12:23:01Z",
  "url": "https://research.draconitic.com/paper/fj84gf4b9qtnv",
  "venue": "Proceedings of the National Academy of Sciences of the United States of America",
  "versions": []
}