Chromatopanning for the identification of gallium binding peptides

•Chromatography-based application of phage surface display for trivalent metal ions.•Stable immobilization of gallium ions as a biopanning target.•Highly stringent biopanning for the enrichment of specific bacteriophage clones.•Successful identification of gallium binding bacteriophage clones. This...

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Bibliographic Details
Published in:Journal of Chromatography A 2019-08, Vol.1600, p.158-166
Main Authors: Schönberger, Nora, Braun, Robert, Matys, Sabine, Lederer, Franziska L., Lehmann, Falk, Flemming, Katrin, Pollmann, Katrin
Format: Article
Language:English
Subjects:
Chromatopanning
Gallium
Peptides
Phage display
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Summary:•Chromatography-based application of phage surface display for trivalent metal ions.•Stable immobilization of gallium ions as a biopanning target.•Highly stringent biopanning for the enrichment of specific bacteriophage clones.•Successful identification of gallium binding bacteriophage clones. This study is concerned with a chromatography-based approach (Immobilized Metal Ion Affinity Chromatography) for the recovery of gallium binding peptide sequences from a recombinant phage display library. The here described methods apply the fundamental knowledge and methods of separation science and meet thereby the key requirement of the phage display technique of precise separation of target-binding bacteriophage clones from non-interacting bacteriophage during the biopanning. During the chromatopanning called process, a total of 101 bacteriophage clones were identified of which in subsequent binding experiments, phage clones expressing the peptide sequences TMHHAAIAHPPH, SQALSTSRQDLR and HTQHIQSDDHLA were characterized to bind >10 fold better to a target that presents immobilized gallium ions than control phage, displaying no peptide sequence. The performance of biopanning experiments in chromatographic systems is particularly suitable for demanding targets such as trivalent metal ions. We found, that the selection process benefits immensely from the stable immobilization of the target metal ions during the entire biopanning process as well as the complete recovery of well interacting bacteriophage clones. Among others, this was possible due to an enhanced monitoring of process conditions and fractionation of eluates.
ISSN:0021-9673
DOI:10.1016/j.chroma.2019.04.037