Protein components of the erythromycin binding site in bacterial ribosomes.
Two derivatives of erythromycin have been prepared carrying either an aryl azide or a 4-nitroguaiacol as a photoreactive group. Both derivatives bind to the specific erythromycin ribosomal site as shown by saturation and competition studies. The derivatives were isotopically labeled either with tritium or with 125I, and radioactivity is covalently incorporated to the ribosome upon irradiation at the appropriate wavelength. The ribosomal proteins labeled were identified by either mono- and two-dimensional gel electrophoresis or high performance liquid chromatography. It has been found that protein L22 is the protein mainly, and under some conditions exclusively, labeled by the erythromycin derivatives. These results were confirmed using ribosomes from erythromycin-resistant mutants having a protein L22 with modified electrophoretical mobility. Protein L15 is also labeled in both cases, and the aromatic azide derivative labels to a lesser extent proteins L2 and L4. Competition experiments with erythromycin indicate that labeling in protein L22, and probably in L15, is specific, while the specificity of labeling in proteins L2 and L4 is questionable. These results indicate that the erythromycin derivatives label different ribosomal proteins than the spiramycin type of macrolides (Tejedor, F., and Ballesta, J.P.G. (1985) Biochemistry 24, 467) suggesting that the binding sites of both macrolide types are probably not identical.
Arévalo
M A
MA
Centro de Biologia Molecular, Consejo Superior de Investigaciones Cientificas-Universidad Autónoma, Madrid, Spain.
Tejedor
F
F
Polo
F
F
Ballesta
J P
JP
D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004917:Erythromycin / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D007700:Kinetics; D011485:Protein Binding; D012269:Ribosomal Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
D012269:Ribosomal Proteins; D004917:Erythromycin