De novo designed 3-helix bundle peptides and proteins with controlled topology and stability Article Swipe
Xiyue Leng
,
Katherine I. Albanese
,
L. Golub
,
Arthur A Norman
,
Jonathan Clayden
,
Derek N. Woolfson
·
YOU?
·
· 2025
· Open Access
·
· DOI: https://doi.org/10.1101/2025.06.30.662276
YOU?
·
· 2025
· Open Access
·
· DOI: https://doi.org/10.1101/2025.06.30.662276
Computational protein design is advancing rapidly. However, approaches and methods are needed to increase success rates and to elaborate designs. Here we describe the combination of rational and computational design to deliver three-helix bundle (3HB) peptide assemblies and single-chain proteins with control over topology and thermal stability. First, we garner sequence-to-structure relationships from antiparallel 3HBs in the Protein Data Bank. This gives core-packing rules, including layers of hydrogen-bonded polar residues, which are combined with surface-charge patterning to design complementary sequences for acidic (A), basic (B), and neutral (N) helices. By altering the design of the N helix, two sets of synthetic peptides are generated for clockwise and anticlockwise arrangements of the three-helix assemblies. Solution-phase characterization shows that both ABN peptide mixtures form stable, heterotrimeric assemblies consistent with the targeted ‘up-down-up’ topologies. Next, AlphaFold2 models for both designs are used to seed computational designs of single-chain proteins where the helices are connected by loop building. Synthetic genes for these express in E. coli to yield soluble, monomeric, and thermally stable proteins. By systematically introducing additional polar layers within the core, the thermal stability of these proteins is varied without compromising the specificity of the helix-helix interactions. Chemical and thermal denaturation reveals comparable thermodynamic parameters to those of highly stable natural proteins. Four X-ray crystal structures confirm that the design models and AlphaFold2 predictions match to sub-Å accuracy.
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Metadata
- Type
- preprint
- Language
- en
- Landing Page
- https://doi.org/10.1101/2025.06.30.662276
- https://www.biorxiv.org/content/biorxiv/early/2025/07/01/2025.06.30.662276.full.pdf
- OA Status
- green
- References
- 75
- Related Works
- 10
- OpenAlex ID
- https://openalex.org/W4411869506
All OpenAlex metadata
Raw OpenAlex JSON
- OpenAlex ID
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https://openalex.org/W4411869506Canonical identifier for this work in OpenAlex
- DOI
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https://doi.org/10.1101/2025.06.30.662276Digital Object Identifier
- Title
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De novo designed 3-helix bundle peptides and proteins with controlled topology and stabilityWork title
- Type
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preprintOpenAlex work type
- Language
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enPrimary language
- Publication year
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2025Year of publication
- Publication date
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2025-07-01Full publication date if available
- Authors
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Xiyue Leng, Katherine I. Albanese, L. Golub, Arthur A Norman, Jonathan Clayden, Derek N. WoolfsonList of authors in order
- Landing page
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https://doi.org/10.1101/2025.06.30.662276Publisher landing page
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https://www.biorxiv.org/content/biorxiv/early/2025/07/01/2025.06.30.662276.full.pdfDirect link to full text PDF
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YesWhether a free full text is available
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greenOpen access status per OpenAlex
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https://www.biorxiv.org/content/biorxiv/early/2025/07/01/2025.06.30.662276.full.pdfDirect OA link when available
- Concepts
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Helix bundle, Bundle, Topology (electrical circuits), Helix (gastropod), Stability (learning theory), Computer science, Chemistry, Biology, Mathematics, Protein structure, Materials science, Biochemistry, Combinatorics, Snail, Machine learning, Composite material, EcologyTop concepts (fields/topics) attached by OpenAlex
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0Total citation count in OpenAlex
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75Number of works referenced by this work
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10Other works algorithmically related by OpenAlex
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| referenced_works | https://openalex.org/W4391431979, https://openalex.org/W4309485183, https://openalex.org/W2351971202, https://openalex.org/W3184152800, https://openalex.org/W2519539312, https://openalex.org/W3137550543, https://openalex.org/W3005709543, https://openalex.org/W2130479394, https://openalex.org/W4404797546, https://openalex.org/W2621783632, https://openalex.org/W2201713963, https://openalex.org/W2968494487, https://openalex.org/W4383957026, https://openalex.org/W4392544734, https://openalex.org/W4321480048, https://openalex.org/W4408934460, https://openalex.org/W4399581345, https://openalex.org/W4323276421, https://openalex.org/W2553669656, https://openalex.org/W2114598719, https://openalex.org/W2138323709, https://openalex.org/W2345914382, https://openalex.org/W3156658374, https://openalex.org/W2465773022, https://openalex.org/W4406387881, https://openalex.org/W4391547180, https://openalex.org/W2518392734, https://openalex.org/W4408246576, https://openalex.org/W2038573488, https://openalex.org/W4285386073, https://openalex.org/W4400356615, https://openalex.org/W4296502374, https://openalex.org/W4399861354, https://openalex.org/W2007606609, https://openalex.org/W2020921026, https://openalex.org/W2071024744, https://openalex.org/W2084376937, https://openalex.org/W2015351966, https://openalex.org/W2961679963, https://openalex.org/W2050888116, https://openalex.org/W2124309900, https://openalex.org/W2027736549, https://openalex.org/W2038051119, https://openalex.org/W4311902557, https://openalex.org/W4387115633, https://openalex.org/W1989504189, https://openalex.org/W3001204511, https://openalex.org/W4409430525, https://openalex.org/W1971470295, https://openalex.org/W2043829693, https://openalex.org/W2131612682, https://openalex.org/W2474147648, https://openalex.org/W3017899883, https://openalex.org/W2126570516, https://openalex.org/W3198143332, https://openalex.org/W2008708467, https://openalex.org/W4255594044, https://openalex.org/W2044698245, https://openalex.org/W2389047281, https://openalex.org/W1998390497, https://openalex.org/W4211239749, https://openalex.org/W2116076986, https://openalex.org/W1989297730, https://openalex.org/W2001458612, https://openalex.org/W3105751341, https://openalex.org/W1495475575, https://openalex.org/W1524706855, https://openalex.org/W2046832917, https://openalex.org/W2746630288, https://openalex.org/W2617643277, https://openalex.org/W4384820649, https://openalex.org/W2735621019, https://openalex.org/W1968673134, https://openalex.org/W2886635500, https://openalex.org/W2032281682 |
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