Insights into the mechanism of azithromycin interaction with an Escherichia coli functional ribosomal complex.
Azithromycin, a derivative of erythromycin with improved activity against Gram-negative bacteria, exhibits a marginal inhibition effect in a model system derived from Escherichia coli, in which a peptide bond is formed between puromycin and AcPhe-tRNA bound at the P-site of poly(U)-programmed ribosomes. This renders the study of azithromycin interaction with Ac[(3)H]Phe-tRNA. poly(U). 70S ribosome complex (complex C) impossible, if we analyze its effect on peptide bond formation. To overcome this problem, we have used an alternative approach to investigate kinetically the azithromycin interaction with complex C and to compare the azithromycin binding properties with those of erythromycin. This approach was based on the ability of azithromycin to compete with tylosin, a macrolide antibiotic strongly inhibiting the puromycin reaction. Detailed kinetic analysis revealed that the encounter complex CA between complex C and azithromycin (A) undergoes a slow isomerization to a tighter complex C*A, which remains active toward puromycin. The determination of inhibition and isomerization rate constants enabled us to classify azithromycin as a slow-binding ligand of ribosomes. Compared with erythromycin, azithromycin is a better inducer and stabilizer of the C*A complex. This finding may explain the superiority of azithromycin as inhibitor of translation in E. coli cells and many other Gram-negative bacteria.
Dinos
G P
GP
Laboratory of Biochemistry, School of Medicine, University of Patras, GR-26500 Patras, Greece.
Michelinaki
M
M
Kalpaxis
D L
DL
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D017963:Azithromycin / Q000494:pharmacology*; D004347:Drug Interactions; D004926:Escherichia coli / Q000187:drug effects* / Q000378:metabolism; D007700:Kinetics; D012270:Ribosomes / Q000187:drug effects* / Q000378:metabolism; D015645:Tylosin / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D017963:Azithromycin; D015645:Tylosin