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Cited by 3Protein Structure and DynamicsRNA and protein synthesis mechanismsOrigins and Evolution of Life

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Folding and knotting of biotic and prebiotic amino acid sequences through reverse evolution

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Especial, João N.C., Faísca, Patrícia F.N.·2025·Biophysical Journal
DOI 10.1016/j.bpj.2025.07.030

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Abstract

We developed a simple reverse evolution method to explore the protein folding transition and knotting process in globular proteins throughout evolution using as a proxy for evolutionary time the length of the amino acid alphabet. Three small proteins were considered. An unknotted one featuring a β-sandwich fold (FN3), a protein embedding a shallow trefoil knot (MJ0366), and a deeply knotted trefoil protein (YibK). Results from Monte Carlo simulations of a native-centric C α model show that thermal stability increases throughout evolution for all considered proteins suggesting it provided a selective pressure for the introduction of biosynthetic amino acids in the alphabet repertoire. Additionally, the thermodynamic cooperativity, the two-state nature of the folding transition, and the kinetic efficiency of folding (and knotting) displayed by the unknotted (and shallow knotted) protein have been roughly conserved throughout evolution, indicating that early alphabets, and, in particular the putative ancestral one composed of 10 amino acids, is competent to self-assemble relatively complex native structures. However, while the ancestral alphabet is clearly more efficient in collapsing the polypeptide chain to nonspecific (i.e., non-native) knots, it significantly compromises the thermodynamic cooperativity and the folding ability of the deeply knotted protein YibK. The findings presented here collectively suggest that the alphabet of amino acids evolved to improve folding, but early alphabets might not have favored the folding of native structures with deep knots. This could help to explain why deep knots in proteins are statistically uncommon.

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Metadata

Authors
Especial, J. N. C., & Faísca, P. F. N.
Affiliations
University of Lisbon (PT)
Corresponding author
Faísca, Patrícia F.N.
ORCID
Venue
Biophysical Journal
Publisher
Elsevier BV
Published
2025-09
Volume
124
Issue
17
Pages
2877-2887
ISSN
0006-3495 (ISSN-L)
DOI
10.1016/j.bpj.2025.07.030
OpenAlex
W4412785468
Cited by
3
References
2
Citations by year
2026: 3
FWCI
0.83
Type
journal article
Language
en
Open access
Licence
Locations
  • Biophysical Journal (publishedVersion)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • PubMed Central (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • University of the East (LISBOA-01-0145-FEDER-022153, 2022.26279, CPCA.A0)
  • European Commission (GA 101079147)
  • Foundation for Science and Technology (SFRH/BD/144345/2019)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
  • Physical Sciences › Physics and Astronomy › Astronomy and Astrophysics › Origins and Evolution of Life
Keywords
protein folding, reverse evolution, prebiotic amino acids, amino acid selection, two-state folding, folding kinetics, protein thermal stability
MeSH terms
Amino Acid Sequence, Kinetics, Models, Molecular, Monte Carlo Method, Proteins, Thermodynamics, Protein Folding, Evolution, Molecular

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CoChat. (2026). Citation check: Folding and knotting of biotic and prebiotic amino acid sequences through reverse evolution. https://app.cochat.ai/papers/10.1016/j.bpj.2025.07.030
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Especial, J. N. C., & Faísca, P. F. N. (2025). Folding and knotting of biotic and prebiotic amino acid sequences through reverse evolution. Biophysical Journal. https://doi.org/10.1016/j.bpj.2025.07.030