Adhiron: a stable and versatile peptide display scaffold for molecular recognition applications.

Journal article

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.

Authors: Tiede, C; Tang, A; Deacon, SE; Mandal, U; Nettleship, J

• Publication status: Published
• Publisher: Oxford University Press
• Journal: Protein engineering, design and selection : PEDS
• Volume: 27
• Issue: 5
• Pages: 145-155
• Publication date: 2014-05-01
• DOI: 10.1093/protein/gzu007
• EISSN: 1741-0134
• ISSN: 1741-0126
• Language: English
• Keywords: non-antibody-binding protein; protein–protein interaction; SUMO; high specificity binding; consensus protein