CHANGES IN RIBOSOMAL ACTIVITY OF ESCHERICHIA-COLI-CELLS DURING PROLONGED CULTURE IN SEA SALTS MEDIUM

The activity of ribosomes from a clinical isolate of Escherichia coli, exposed to starvation for 7 days ire sea salts medium, was investigat ed by measuring the kinetic parameters of ribosomal peptidyltransferas e, by using the puromycin reaction as a model reaction. No alterations in the extent of peptide bond formation were observed during starvati on. In contrast, a 50% reduction in the k(max)/K-s ratio could he seen after 24 h of starvation; an additional 6 days of starvation resulted in a progressive but less abrupt decline in the k(max)/K-s value. {k( max) is the: apparent catalytic rate constant of peptidyl transferase, and K-s is the dissociation constant of the encounter complex; betwee n acetyl (Ac)[H-3]Phe-tRNA-poly(U)-ribosome and puromycin.} Although t he distribution of ribosomal particles remained constant, a substantia l decrease in the number of ribosomes per starved cell and a clear dec line in the ability of ribosomes to bind AcPhe-tRNA were observed, par ticularly during the first day of starvation. Further analysis indicat ed that rRNA ire general, hut especially 23S rRNA, was rapidly degrade d during the starvation period. In addition, the L12/L7 molar ratio de creased from 1.5 to a during the initial phase of starvation (anp to 2 4 h) but remained constant during the subsequent starvation period. Ri bosomes isolated from 24-h-starved cells, when artificially depleted o f L7/L12 protein and reconstituted with L7/L12 protein from mid-logari thmic-phase cells, regenerated an L12/L7 molar ratio of 1.5 and restor ed the Peptidyltransferase activity to a substantial level. An analogo us effect of reconstitution on the efficiency of ribosomes in binding AcPhe-tRNA was evident not only during the initial phase hut throughou t the starvation period.