Iwan Zimmermann, Pascal Egloff, Cédric Hutter, Fabian M. Arnold, Peter Stohler, Nicolas Bocquet, Melanie N. Hug, Sylwia Huber, Martin Siegrist, Lisa Hetemann, Jennifer Gera, Samira Gmür, Peter Spies, Daniel Gygax, Eric R. Geertsma, Roger Dawson, Markus A. Seeger
- Mechanistic and structural studies of membrane proteins require their stabilization in specific conformations. Single domain antibodies are potent reagents for this purpose, but their generation relies on immunizations, which impedes selections in the presence of ligands typically needed to populate defined conformational states. To overcome this key limitation, we developed an in vitro selection platform based on synthetic single domain antibodies named sybodies. To target the limited hydrophilic surfaces of membrane proteins, we designed three sybody libraries that exhibit different shapes and moderate hydrophobicity of the randomized surface. A robust binder selection cascade combining ribosome and phage display enabled the generation of conformation-selective, high affinity sybodies against an ABC transporter and two previously intractable human SLC transporters, GlyT1 and ENT1. The platform does not require access to animal facilities and builds exclusively on commercially available reagents, thus enabling every lab to rapidly generate binders against challenging membrane proteins.
MetadatenAuthor: | Iwan Zimmermann, Pascal Egloff, Cédric Hutter, Fabian M. Arnold, Peter Stohler, Nicolas Bocquet, Melanie N. Hug, Sylwia Huber, Martin Siegrist, Lisa Hetemann, Jennifer Gera, Samira Gmür, Peter Spies, Daniel Gygax, Eric R. GeertsmaORCiD, Roger Dawson, Markus A. SeegerORCiD |
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URN: | urn:nbn:de:hebis:30:3-503554 |
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DOI: | https://doi.org/10.7554/eLife.34317.001 |
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ISSN: | 2050-084X |
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Pubmed Id: | https://pubmed.ncbi.nlm.nih.gov/29792401 |
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Parent Title (English): | eLife |
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Publisher: | eLife Sciences Publications |
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Place of publication: | Cambridge |
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Contributor(s): | Lucy Forrest |
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Document Type: | Article |
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Language: | English |
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Year of Completion: | 2018 |
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Date of first Publication: | 2018/05/24 |
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Publishing Institution: | Universitätsbibliothek Johann Christian Senckenberg |
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Release Date: | 2019/06/13 |
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Tag: | Biochemistry and chemical biology; Conformational trapping; E. colo; In vitro selection; Membrane protein; Nanobody; Phage display; Ribosome display; Structural biology and molecular biophysics; Tools and ressources |
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Volume: | 7 |
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Issue: | e34317 |
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Page Number: | 32 |
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First Page: | 1 |
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Last Page: | 32 |
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Note: | Copyright Zimmermann et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited. |
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HeBIS-PPN: | 450752313 |
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Institutes: | Biochemie, Chemie und Pharmazie / Biochemie und Chemie |
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Dewey Decimal Classification: | 5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie |
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Sammlungen: | Universitätspublikationen |
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Licence (German): | Creative Commons - Namensnennung 4.0 |
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