Identification of antibiotic clarithromycin binding peptide displayed by T7 phage particles.
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 2.1 x 10(-3) M. The dissociation constants of other macrolide antibiotics, erythromycin, roxithromycin, azithromycin and josamycin were also determined to be 5.4 x 10(-3) M, 6.2 x 10(-5) M, 1.1 M and 3.4 x 10(-2) 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.
Morimura
Tetsuro
T
Genome and Drug Research Center, Department of Applied Biological Science, Tokyo University of Science, Noda, Chiba 278-8510, Japan.
Noda
Naoko
N
Kato
Yasutaro
Y
Watanabe
Tetsuaki
T
Saitoh
Takeki
T
Yamazaki
Takayuki
T
Takada
Keiichi
K
Aoki
Satoko
S
Ohta
Keisuke
K
Ohshige
Masahiko
M
Sakaguchi
Kengo
K
Sugawara
Fumio
F
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000378:metabolism*; D017123:Bacteriophage T7 / Q000235:genetics*; D019921:Biotinylation; D017291:Clarithromycin / Q000031:analogs & derivatives / Q000378:metabolism*; D015723:Gene Library; D008024:Ligands; D008969:Molecular Sequence Data; D015394:Molecular Structure; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D020349:Surface Plasmon Resonance
D000900:Anti-Bacterial Agents; D008024:Ligands; D019151:Peptide Library; D010455:Peptides; D017291:Clarithromycin