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Efficient one-cycle affinity selection of binding proteins or peptides specific for a small-molecule using a T7 phage display pool by Yoichi Takakusagi; Kouji Kuramochi; Manami Takagi; Tomoe Kusayanagi; Daisuke Manita; Hiroko Ozawa; Kanako Iwakiri; Kaori Takakusagi; Yuka Miyano; Atsuo Nakazaki; Susumu Kobayashi; Fumio Sugawara; Kengo Sakaguchi is a Biochemistry, Genetics and Molecular Biology article available to read on EtoBox.

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Here, we report an efficient one-cycle affinity selection using a natural-protein or random-peptide T7 phage pool for identification of binding proteins or peptides specific for small-molecules. The screening procedure involved a cuvette type 27-MHz quartz-crystal microbalance (QCM) apparatus with introduction of self-assembled monolayer (SAM) for a specific small-molecule immobilization on the gold electrode surface of a sensor chip. Using this apparatus, we attempted an affinity selection of proteins or peptides against synthetic ligand for FK506-binding protein (SLF) or irinotecan (Iri, CPT-11). An affinity selection using SLF-SAM and a natural-protein T7 phage pool successfully detected FK506-binding protein 12 (FKBP12)-displaying T7 phage after an interaction time of only 10 min. Extensive exploration of time-consuming wash and/or elution conditions together with several rounds of selection was not required. Furthermore, in the selection using a 15-mer random-peptide T7 phage pool and subsequent analysis utilizing receptor ligand contact (RELIC) software, a subset of SLF-selected peptides clearly pinpointed several amino-acid residues within the binding site of FKBP12. Likewis

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Author

Yoichi Takakusagi; Kouji Kuramochi; Manami Takagi; Tomoe Kusayanagi; Daisuke Manita; Hiroko Ozawa; Kanako Iwakiri; Kaori Takakusagi; Yuka Miyano; Atsuo Nakazaki; Susumu Kobayashi; Fumio Sugawara; Kengo Sakaguchi

Publisher

Elsevier Science; Elsevier ; Elsevier Ltd.; Elsevier BV (ISSN 0968-0896)

Published

2008

Language

EN

Field

Biochemistry, Genetics and Molecular Biology (Life Sciences)