Assembly and use of high-density recombinant peptide chips for large-scale ligand screening is a practical alternative to synthetic peptide libraries

BMC Genomics

Methodology Article

Harald Hundsberger¹  Dezso P. Virok²  Julia Herzog³  Raphaela Rid³  Kamil Önder⁴  Peter Schuller-Götzburg⁵ 

[1] Department of Medical and Pharmaceutical Biotechnology, University of Applied Sciences, 3500, Krems, Austria;Department of Medicinal Microbiology and Immunobiology, University of Szeged, 6722, Szeged, Hungary;Research Program for Rational Drug Design in Dermatology and Rheumatology, Department of Dermatology, Paracelsus Medical University of Salzburg, 5020, Salzburg, Austria;Research Program for Rational Drug Design in Dermatology and Rheumatology, Department of Dermatology, Paracelsus Medical University of Salzburg, 5020, Salzburg, Austria;ProComCure Biotech, 5081, Anif, Austria;Research Program in Prosthetics, Biomechanics and Biomaterials, Paracelsus Private Medical University, 5020, Salzburg, Austria;

关键词: Peptide library;    Recombinational cloning;    Recombinant peptide array;    Protein chip;    Peptide screening;    Protein tags;    Ligand design;    High affinity;    Target binding;    Diversity;   

DOI  :  10.1186/s12864-017-3814-3

 received in 2017-02-14, accepted in 2017-05-23,  发布年份 2017

来源: Springer

【 摘 要 】

BackgroundRecombinant peptide chips could constitute a versatile complementation to state-of-the-art in situ (chemical on-chip) synthesis, particle-based printing, or pre-manufactured peptide spotting. Bottlenecks still impeding a routine implementation - from restricted peptide lengths, low diversity and low array densities to high costs - could so be overcome.MethodsTo assess overall performance, we assembled recombinant chips composed of 38,400 individual peptide spots on the area of a standard 96-well microtiter plate from comprehensive, highly diverse (>107 single clones) short random peptide libraries.ResultsScreening of altogether 476,160 clones against Streptavidin uncovered 2 discrete new binders: a characteristic HPQ-motif containing VSHPQAPF and a cyclic CSGSYGSC peptide. Interactions were technically confirmed by fluorescence polarization as well as biolayer-interferometry, and their potential suitability as novel detection tags evaluated by detection of a peptide-fused exemplary test protein.ConclusionFrom our data we conclude that the presented technical pipeline can reliably identify novel hits, useful as first-generation binders or templates for subsequent ligand design plus engineering.

【 授权许可】

CC BY   
© The Author(s). 2017