Mechanism of action of gentamicin components

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Abstract
The binding of gentamicin (Gm) to E. coli ribosomes and ribosomal subunits was studied. By means of equilibrium dialysis and of statistical interpretation of the data it was found that [3H]gentamicin C2 and 6''-N-[3H]methylgentamicin C1a interact with 3 classes of sites on tight-coupled 70S species: a 1st class concerning the tight and non-cooperative interaction with 1 drug molecule (Kd = 0.6 .mu.M), a 2nd class in which .apprx. 5 Gm molecules bind cooperatively (mean Kd = 10 .mu.M) and a 3rd class of very high capacity in which up to 70 drug molecules may interact. The extreme cooperativity of the 3rd class of sites induces such an increase in the affinity for Gm that it may allow the shift of molecules already bound from high-affinity sites towards lower-affinity sites. The alteration of a ribosomal protein, L6, in a Gm-resistant mutant of E. coli abolished the multiclass and the cooperative aspects of ribosomes-Gm interaction. The large ribosomal subunits from E. coli MRE 600 strain interact cooperatively with Gm, whereas 50S particles from the resistant mutant bind the drug in a diffuse way with high capacity and low affinity. The small subunits from both strains behave identically towards Gm. A good correlation is observed in comparing the Gm concentration capable of saturating the different ribosomal classes of sites with concentrations inducing its multiphasic effects on protein synthesis.