Tiede, C orcid.org/0000-0003-0280-4005, Tang, AA orcid.org/0000-0002-5216-5080, Deacon, SE et al. (8 more authors) (2014) Adhiron: a stable and versatile peptide display scaffold for molecular recognition applications. Protein Engineering, Design and Selection, 27 (5). pp. 145-155. ISSN 1741-0126
Abstract
We have designed a novel non-antibody scaffold protein, termed Adhiron, based on a phytocystatin consensus sequence. The Adhiron scaffold shows high thermal stability (Tm ca. 101°C), and is expressed well in Escherichia coli. We have determined the X-ray crystal structure of the Adhiron scaffold to 1.75 Å resolution revealing a compact cystatin-like fold. We have constructed a phage-display library in this scaffold by insertion of two variable peptide regions. The library is of high quality and complexity comprising 1.3 × 10(10) clones. To demonstrate library efficacy, we screened against the yeast Small Ubiquitin-like Modifier (SUMO). In selected clones, variable region 1 often contained sequences homologous to the known SUMO interactive motif (V/I-X-V/I-V/I). Four Adhirons were further characterised and displayed low nanomolar affinities and high specificity for yeast SUMO with essentially no cross-reactivity to human SUMO protein isoforms. We have identified binders against >100 target molecules to date including as examples, a fibroblast growth factor (FGF1), platelet endothelial cell adhesion molecule (PECAM-1; CD31), the SH2 domain Grb2 and a 12-aa peptide. Adhirons are highly stable and well expressed allowing highly specific binding reagents to be selected for use in molecular recognition applications.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © The Author 2014. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. |
Keywords: | SUMO; consensus protein; high specificity binding; non-antibody-binding protein; protein–protein interaction |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) > Synthetic Biology (Leed) The University of Leeds > Faculty of Medicine and Health (Leeds) > School of Medicine (Leeds) > Inst of Clinical Trials Research (LICTR) (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 18 Dec 2014 12:52 |
Last Modified: | 20 Jun 2021 08:38 |
Published Version: | http://dx.doi.org/10.1093/protein/gzu007 |
Status: | Published |
Publisher: | Oxford University Press |
Refereed: | Yes |
Identification Number: | 10.1093/protein/gzu007 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:81756 |