Persistent Identifier
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doi:10.11588/data/SGN2CN |
Publication Date
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2023-02-10 |
Title
| Identification of novel functional mini-receptors by combinatorial screening of split-WW domains [Research Data] |
Author
| Thomas, Franziska (Heidelberg University, Institute of Organic Chemistry) - ORCID: 0000-0002-1176-7018
Neitz, Hermann (University of Würzburg, Institute of Organic Chemistry) - ORCID: 0000-0001-9088-5727
Paul, Niels Benjamin (University of Göttingen, Institute of Organic and Biomolecular Chemistry) - ORCID: 0000-0002-5631-0126
Häge, Florian (Heidelberg University, Institute of Organic Chemistry) - ORCID: 0000-0003-4055-1168
Lindner, Christina (Heidelberg University, Institute of Organic Chemistry) - ORCID: 0000-0003-2910-3022
Graebner, Roman (Heidelberg University, Institute of Organic Chemistry) - ORCID: 0000-0001-8568-3485
Kovermann, Michael (University of Konstanz, Department of Chemistry) - ORCID: 0000-0002-3357-9843 |
Point of Contact
|
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Thomas, Franziska (Heidelberg University, Institute of Organic Chemistry)
Kovermann, Michael (University of Konstanz, Department of Chemistry) |
Description
| β-Sheet motifs such as the WW domain are increasingly being explored as building blocks for synthetic biological applications. Since the sequence-structure relationships of β-sheet motifs are generally complex compared to the well-studied α-helical coiled coil (CC), other approaches such as combinatorial screening should be included to vary the function of the peptide. In this study, we present a combinatorial approach to identify novel functional mini-proteins based on the WW-domain scaffold, which takes advantage of the successful reconstitution of the fragmented WW domain of hPin1 (hPin1WW) by CC association. Fragmentation of hPin1WW was performed in both loop 1 (CC-hPin1WW-L1) and loop 2 (CC-hPin1WW-L2), and the respective fragments were linked to the strands of an antiparallel heterodimeric CC. Structural analysis by CD and NMR spectroscopy revealed structural reconstitution of the WW-domain scaffold only in CC-hPin1WW-L1, but not in CC-hPin1WW-L2. Furthermore, by using 1H–15N HSQC NMR, fluorescence and CD spectroscopy, we demonstrated that binding properties of fragmented hPin1WW in CC-hPin1WW-L1 were fully restored by CC association. To demonstrate the power of this approach as a combinatorial screening platform, we synthesized a four-by-six library of N- and C-terminal hPin1WW-CC peptide fragments that was screened for a WW domain that preferentially binds to ATP over cAMP, phophocholine, or IP6. Using this screening platform, we identified one WW domain, which specifically binds ATP, and a phosphorylcholine-specific WW-based mini-receptor, both having binding dissociation constants in the lower micromolar range. |
Subject
| Chemistry |
Keyword
| miniprotein
WW domain
coiled coil
peptide engineering |
Related Publication
| Neitz, H., Paul, N. B., Häge, F., Lindner, C., Graebner, R., Kovermann, M., & Thomas, F. (2022). Identification of novel functional mini‐receptors by combinatorial screening of split‐WW domains. Chemical Science, 13, 9079–9090. doi: 10.1039/D2SC01078J https://doi.org/10.1039/D2SC01078J |
Funding Information
| Deutsche Forschungsgemeinschaft (DFG): 414261058, 2082/1–390761711 (3DMM2O)
Young Scholar Fund: (M.K)
University of Konstanz: (M.K)
Carl Zeiss Foundation: (F.H)
Dr Sophie-Bernthsen-Stiftung |