Fluorescence polarization method to characterize macrolide-ribosome interactions.
A fluorescence polarization assay is described that measures the binding of fluorescently labeled erythromycin to 70S ribosomes from Escherichia coli and the displacement of the erythromycin from these ribosomes. The assay has been validated with several macrolide derivatives and other known antibiotics. We demonstrate that this assay is suitable for determining the dissociation constants of novel compounds that have binding sites overlapping those of macrolides. This homogeneous binding assay provides a valuable tool for defining structure-activity relationships among compounds during the discovery and development of new ribosome-targeting drugs.
Yan
Kang
K
1250 South Collegeville Road, Collegeville, PA 19426, USA. kang.2.yan@gsk.com
Hunt
Eric
E
Berge
John
J
May
Earl
E
Copeland
Robert A
RA
Gontarek
Richard R
RR
D000900:Anti-Bacterial Agents / Q000378:metabolism*; D001665:Binding Sites; D004917:Erythromycin / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D005454:Fluorescence Polarization / Q000379:methods*; D018942:Macrolides / Q000378:metabolism*; D012335:RNA, Ribosomal / Q000235:genetics; D012270:Ribosomes / Q000737:chemistry / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D018942:Macrolides; D012335:RNA, Ribosomal; D004917:Erythromycin