Polyamines affect diversely the antibiotic potency: insight gained from kinetic studies of the blasticidin S AND spiramycin interactions with functional ribosomes.
The effects of spermine on peptidyltransferase inhibition by an aminohexosylcytosine nucleoside, blasticidin S, and by a macrolide, spiramycin, were investigated 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. Kinetics revealed that blasticidin S, after a transient phase of interference with the A-site, is slowly accommodated near to the P-site so that peptide bond is still formed but with a lower catalytic rate constant. At high concentrations of blasticidin S (>10 x K(i)), a second drug molecule binds to a weaker binding site on ribosomes, and this may account for the onset of a subsequent mixed-noncompetitive inhibition phase. Spermine enhances the blasticidin S inhibitory effect by facilitating the drug accommodation to both sites. On the other hand, spiramycin (A) was found competing with puromycin for the A-site of AcPhe-tRNA.poly(U).70 S ribosomal complex (C) via a two-step mechanism, according to which the fast formation of the encounter complex CA is followed by a slow isomerization to a tighter complex, termed C(*)A. In contrast to that observed with blasticidin S, spermine reduced spiramycin potency by decreasing the formation and stability of complex C(*)A. Polyamine effects on drug binding were more pronounced when a mixture of spermine and spermidine was used, instead of spermine alone. Our kinetic results correlate well with cross-linking and crystallographic data and suggest that polyamines bound at the vicinity of the antibiotic binding pockets modulate diversely the interaction of these drugs with ribosomes.
Petropoulos
Alexandros D
AD
Laboratory of Biochemistry, School of Medicine, University of Patras, GR-26500 Patras, Greece.
Xaplanteri
Maria A
MA
Dinos
George P
GP
Wilson
Daniel N
DN
Kalpaxis
Dimitrios L
DL
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D004926:Escherichia coli / Q000378:metabolism; D007700:Kinetics; D008954:Models, Biological; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009705:Nucleosides / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D011073:Polyamines / Q000737:chemistry*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011691:Puromycin / Q000737:chemistry; D012338:RNA, Ribosomal, 23S / Q000737:chemistry; D012270:Ribosomes / Q000378:metabolism*; D015572:Spiramycin / Q000737:chemistry*; D013997:Time Factors
D000900:Anti-Bacterial Agents; D009705:Nucleosides; D010455:Peptides; D011073:Polyamines; D012338:RNA, Ribosomal, 23S; D011691:Puromycin; D015572:Spiramycin; C004500:blasticidin S