Fluorescein and Rhodamine B-Binding Domains from Autodisplayed Fv-Antibody Library

In this study, the binding domains for fluorescent dyes were presented that could be used as synthetic peptides or fusion proteins. Fv-antibodies against two fluorescent dyes (fluorescein and rhodamine B) were screened from the Fv-antibody library, which was prepared on the outer membrane of Escherichia coli using the autodisplay technology. Two clones with binding activities to each fluorescent dye were screened separately from the library using flow cytometry. The binding activity of the screened Fv-antibodies on the outer membrane was analyzed using fluorescent imaging with the corresponding fluorescent dyes. The CDR3 regions of the screened Fv-antibodies (11 amino acid residues) were synthesized into peptides, and each peptide was analyzed for its binding activity to each fluorescent dye using fluorescence resonance energy transfer (FRET) experiments. These CDR3 regions were demonstrated to have a binding activity to each fluorescent dye when the regions were co-expressed as a fusion protein with Z-domain.