Clarithromycin is a macrolide antibiotic that is widely used in clinical medicine. Macrolide antibiotics such as clarithromycin specifically bind to the 50S subunit of the bacterial ribosome thereby interfering with protein biosynthesis. A selected peptide sequence from our former study, composed of 19 amino acids, which was isolated from a phage display library because of its ability to bind clarithromycin, displayed significant similarity to a portion of the human_p8 protein. The recombinant p8 protein binds to biotinylated-clarithromycin immobilized on a streptavidin-coated sensor chip and the dissociation constant was determined. The binding of recombinant p8 protein to double-stranded DNA was inhibited by biotinylated-clarithromycin, clarithromycin, erythromycin and azithromycin in gel mobility shift assay. Dechlorogriseofulvin, obtained from a natural product screening, also inhibited human p8 protein binding to DNA. This study illustrates the general utility of the phage display method in detecting protein-ligand interactions.

Peptide libraries displayed by T7 phage, which contain random cDNA fragments insets, were screened for their ability to bind to a biotinylated derivative of clarithromycin. Phage particles bound to an immobilized derivative of the antibiotic were isolated and the inserted cDNA was amplified and sequenced. A common selected peptide sequence, composed of 19 amino acids, was obtained and a synthetic peptide with this sequence was produced. Surface plasmon resonance experiments showed that the synthetic peptide immobilized on a sensor chip bound to clarithromycin and the dissociation constant was determined to be 2.1ϫ10 Ϫ3 M. The dissociation constants of other macrolide antibiotics, erythromycin, roxithromycin, azithromycin and josamycin were also determined to be 5.4ϫ10 Ϫ3 M, 6.2ϫ10 Ϫ5 M, 1.1 M and 3.4ϫ10 Ϫ2 M, respectively. These results indicated that T7 phage display method might be useful to determine relatively weak interactions between small molecule drugs and the selected peptides which could represent a possible binding site conserved in binding proteins.