Kinetic properties of Escherichia coli ribosomes with altered forms of S12. |
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Authors: | N Bilgin F Claesens H Pahverk M Ehrenberg |
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Affiliation: | University of Uppsala, Department of Molecular Biology, Sweden. |
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Abstract: | E. coli ribosomes with alterations in S12 leading to streptomycin resistance (SmR), dependence (SmD) and pseudodependence (SmP) were studied with the quench-flow technique. Kinetic changes at the various steps of the elongation cycle were identified. The rate of hydrolysis of GTP in the ternary complex in the ribosomal A-site is decreased drastically in SmD and moderately in SmP in relation to wild-type ribosomes. Addition of streptomycin restores much of the wild-type behaviour. The SmD, SmP and SmR ribosomes have an enhanced GTP-hydrolysis idling reaction on EF-Tu, which is correlated with how aggressive proofreaders these ribosomes are in steady-state assays. We use our in vitro findings to discuss the in vivo physiology of these mutants as well as mechanistic features of E. coli translation. |
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