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Cited by 0Protein Structure and DynamicsEnzyme Structure and FunctionGenomics and Phylogenetic Studies

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Tracing the function expansion for a primordial protein fold in the era of fold-based function prediction: β-trefoil

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Goswami, Moushmi, Srinivasan, Subhashini·2025·PLOS One
DOI 10.1371/journal.pone.0320177PMC PMC12225799

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Abstract

The incredibly narrow protein fold bottleneck, which separates the billions of unique proteins on one side to deliver diverse biological functions on the other, arises from folds that tolerate mutations during evolution. One such fold, called the β-trefoil, is present in functionally diverse proteins including cytokines involved in the immune system such as interleukin-1. The unrecognizable sequence-level diversity, even among paralogs of interleukin-1 within the same chromosomal locus, suggests the resilience of this fold to mutational onslaught. Furthermore, β-trefoil domain containing-proteins are known to coexist with other domains to achieve functional diversity. In this study, we challenge the reach and limitations of function prediction using fold-fold comparison using β-trefoil fold as an example. We identified proteins containing β-trefoil fold belonging to thirty-two distinct functional classes based on diverse domain architecture and/or functional annotation by mining both the PDB and AlphaFold databases using fold-fold comparison. Among the proteins with novel domain architecture we find β-trefoil along with chitinase, lipase, β-glucosidase, protein kinase, peptidoglycan-binding + peptidase matrixin, glycosyl hydrolases family 3 + PA14 + fibronectin type- III, alpha galactosidase A, PhoD-like phosphatase, insecticidal crystal toxin, trypsin, alginate lyase and two novel structurally uncharacterized domains. We demonstrate that fold-fold comparison can extend function prediction beyond the reach of sequence-based approach and provides an opportunity to discover novel domain architecture associated with known folds. However, since extending fold similarity to functional similarity may be challenged by convergent fold evolution, we explore if β-trefoil may be a convergent evolution and share our hypothesis.

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Registry

Crossref · 10.1371/journal.pone.0320177

Cites

22 of 22 references verified0 citation errors · 0 not checkable

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Figures

Fig 1 The core of the sundial represents the β-trefoil fold. The second ring classifies the proteins into functional families. The third layer shows the distribution of β-trefoil fold in PDB across biological kingdoms, color-coded as blue for bacteria, gray for fungi, green for plants, red for Metazoa, and yellow for protozoa. Fourth layer shows representative β-hairpin without loops from each functional class along with r.m.s.d. and sequence identity compared to interleukin-1 with the PDBid of 5bow. The outermost layer illustrates domain architecture with purple representing the β-trefoil domain and additional domains in unique colors such as Agglutinin (orange), Glycosyl transferase family (dark-yellow), Glycosyl hydrolases family (dark-blue), Clostridium neurotoxin receptor-binding domain (light-blue), RIH domain (brown), Ryanodine receptor-associated domain (dark-green), Unknown domain 1 (light-pink), Unknown domain 2 (dark-pink), Unknown domain 3 (lemon-yellow), Unknown domain 4 (grey), Unknown domain 5 (pastel-green), Spindlin/Ssty family domain (light-green), Cellulase (red), Carbohydrate binding family domain (forest-green).
Fig 1 The core of the sundial represents the β-trefoil fold. The second ring classifies the proteins into functional families. The third layer shows the distribution of β-trefoil fold in PDB across biological kingdoms, color-coded as blue for bacteria, gray for fungi, green for plants, red for Metazoa, and yellow for protozoa. Fourth layer shows representative β-hairpin without loops from each functional class along with r.m.s.d. and sequence identity compared to interleukin-1 with the PDBid of 5bow. The outermost layer illustrates domain architecture with purple representing the β-trefoil domain and additional domains in unique colors such as Agglutinin (orange), Glycosyl transferase family (dark-yellow), Glycosyl hydrolases family (dark-blue), Clostridium neurotoxin receptor-binding domain (light-blue), RIH domain (brown), Ryanodine receptor-associated domain (dark-green), Unknown domain 1 (light-pink), Unknown domain 2 (dark-pink), Unknown domain 3 (lemon-yellow), Unknown domain 4 (grey), Unknown domain 5 (pastel-green), Spindlin/Ssty family domain (light-green), Cellulase (red), Carbohydrate binding family domain (forest-green).CC BY · PubMed Central
Fig 2 Each plot shows r.m.s.d. (x-axis: 0-3 Å) versus percent identities (y-axis: 30-100) for the eleven selected queries (bold in Table 1 column 2) against AFdb hits with more than 80% query coverage.
Fig 2 Each plot shows r.m.s.d. (x-axis: 0-3 Å) versus percent identities (y-axis: 30-100) for the eleven selected queries (bold in Table 1 column 2) against AFdb hits with more than 80% query coverage.CC BY · PubMed Central
Fig 3 Novel domain architecture of UniProt proteins containing β-trefoil domains predicted by AlphaFold that are not yet represented in the PDB. A tabular format is included in S1 Table in S1 File.
Fig 3 Novel domain architecture of UniProt proteins containing β-trefoil domains predicted by AlphaFold that are not yet represented in the PDB. A tabular format is included in S1 Table in S1 File.CC BY · PubMed Central
Fig 4 Stereo diagram of the six characterized UniProt entries with predicted structures not yet represented in PDB. PDB queries in 4b and 4d representing category-1 are in green, AFdb structures in pink, PDB hits for category-2 in 4a, 4c, 4e and 4f are in blue.
Fig 4 Stereo diagram of the six characterized UniProt entries with predicted structures not yet represented in PDB. PDB queries in 4b and 4d representing category-1 are in green, AFdb structures in pink, PDB hits for category-2 in 4a, 4c, 4e and 4f are in blue.CC BY · PubMed Central
Fig 5 Stereo diagram of the seven uncharacterized proteins from AFdb containing β-trefoil domain not part of the functions and domain architecture presented in Fig 1. PDB queries in 5a and 5b representing category-1 are in green, AFdb structures in orange, PDB hits for categories-2 and 3 in 5c through 5f in blue.
Fig 5 Stereo diagram of the seven uncharacterized proteins from AFdb containing β-trefoil domain not part of the functions and domain architecture presented in Fig 1. PDB queries in 5a and 5b representing category-1 are in green, AFdb structures in orange, PDB hits for categories-2 and 3 in 5c through 5f in blue.CC BY · PubMed Central
Fig 6 Top shows phylogenetic tree from only aligning the 7 core positions with low entropy. Bottom show Shannon entropy at positions with less than 10% gaps from a structure-based alignment. The mutation tolerance at each core position is shown with red arrows.
Fig 6 Top shows phylogenetic tree from only aligning the 7 core positions with low entropy. Bottom show Shannon entropy at positions with less than 10% gaps from a structure-based alignment. The mutation tolerance at each core position is shown with red arrows.CC BY · PubMed Central
Fig 7 Flowchart of various steps along with filtering criteria.
Fig 7 Flowchart of various steps along with filtering criteria.CC BY · PubMed Central

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Metadata

Authors
Goswami, M., & Srinivasan, S.
Affiliations
Institute of Bioinformatics and Applied Biotechnology (IN)
Corresponding author
Srinivasan, Subhashini
ORCID
Srinivasan, Subhashini
Venue
PLOS One
Publisher
Public Library of Science (PLoS)
Published
2025-07-03
Volume
20
Issue
7
Pages
e0320177
ISSN
1932-6203 (ISSN-L)
DOI
10.1371/journal.pone.0320177
PMC
PMC12225799
OpenAlex
W4411995712
Cited by
0
References
22
FWCI
0.00
Type
journal article
Language
en
Open access
Licence
Locations
  • PLoS ONE (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
Funders
Department of Biotechnology, Ministry of Science and Technology, India (BT/PR40212/BTIS/137/40/2022)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Enzyme Structure and Function
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Phylogenetic Studies
Keywords
β-trefoil fold, protein function prediction, domain architecture, interleukin-1, cytokines, convergent evolution, protein fold evolution, protein domains, structural genomics
MeSH terms
Protein Domains, Models, Molecular, Proteins, Protein Folding, Databases, Protein

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