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  • Open Access

Role of topology in protein thermal stability

João N. C. Especial1, Beatriz P. Teixeira2, Ana Nunes1, Miguel Machuqueiro3, and Patrícia F. N. Faísca1,*

  • 1BioISI—Instituto de Biossistemas e Ciências Integrativas and Departamento de Física, Faculdade de Ciências, Universidade de Lisboa, 1749-016 Lisboa, Portugal
  • 2Departamento de Física, Faculdade de Ciências, Universidade de Lisboa, 1749-016 Lisboa, Portugal
  • 3BioISI—Instituto de Biossistemas e Ciências Integrativas and Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade de Lisboa, 1749-016 Lisboa, Portugal

  • *Contact author: pffaisca@ciencias.ulisboa.pt
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Phys. Rev. Research 8, 013284 – Published 13 March, 2026

DOI: https://doi.org/10.1103/9kzy-w2tz

Abstract

For several decades, experimental and computational studies have been used to investigate the potential functional role of knots in protein structures. A property that has attracted considerable attention is thermal stability, i.e., the extent to which a protein retains its native conformation and biological activity at high temperatures, without undergoing denaturation or aggregation. Thermal stability is quantified by the melting temperature Tm, an equilibrium property that corresponds to the peak of heat capacity in differential scanning calorimetry (DSC) experiments. Experimental and computational studies report conflicting effects of knotting on protein thermal stability [M. A. Soler and P. F. N. Faísca, PLoS One 8, e74755 (2013);Y.-C. Chuang, I.-C. Hu, P.-C. Lyu, and S.-T. D. Hsu, J. Mol. Biol. 431, 857 (2019)]. Here, we use extensive Monte Carlo simulations of a simple C-alpha model of protein YibK, with energetics modeled by the Go potential, to show that Tm does not depend on the topological state of the protein. Our simulations further support the view that the discrepancy between the experimental and computational results stems from a pronounced separation of timescales for unknotting and unfolding that is inherent to deeply knotted proteins like YibK. In particular, the timescale separation implies that the complete unfolding-untying transition may not be accessible within the duration of a DSC experiment, whose apparent Tm measurements likely reflect a nonequilibrium distribution lacking unfolded states that are also unknotted.

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Phys. Rev. Research 8, 013284– Published 13 March, 2026
  • Received 10 November 2025
  • Accepted 9 February 2026
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