title: A kirromycin-resistant EF-Tu species reverses streptomycin dependence of Escherichia coli strains mutated in ribosomal protein S12.
abstract: Streptomycin dependence can be caused by mutations in ribosomal protein S12. Mutations suppressing such streptomycin dependence have been found in ribosomal proteins S4 and S5, and in 16S rRNA. Here a new suppressor mutation localized in elongation factor Tu (EF-Tu) is described, consistent with recent models of ribosome-EF-Tu-tRNA interaction at the decoding centre. The EF-Tu mutation was obtained by genetic selection for streptomycin independence; it was identified as Ala375 --> Thr, previously described as EF-TuA(R) and known to confer a kirromycin-resistant, error-prone phenotype. Also, other streptomycin-dependent (SmD) S12 mutations could be complemented by this mutation. The streptomycin-independent (Sm1) strain grows more slowly than the wild-type (wt), suggesting that not all the defects of the S12 mutation can be complemented by EF-Tu[A375T]. Moreover, this strain is more susceptible than wt to reduction in the cellular EF-Tu concentration, and disruption of tufB led to considerable growth-rate impairment. Expression of EF-Tu from tufB, not only of wt EF-Tu and EF-Tu[A375T] but, remarkably, also of EF-Tu[G222D], known as EF-TuB0 and defective in protein synthesis, equally contributed to cell growth. In vitro analysis revealed a decreased translational activity of wt EF-Tu with SmD ribosomes as compared to EF-Tu[A375T], while EF-Tu[G222D] showed no activity at all, just as with wt ribosomes. Possible mechanisms are discussed for the improved growth rate observed in such Sm1 strains when they include wt EF-Tu or EF-Tu[G222D].
authors.forename: Anne-Marie
authors.initials: AM
authors.lastname: Zuurmond
authors.forename: Leo A H
authors.initials: LAH
authors.lastname: Zeef
authors.forename: Barend
authors.initials: B
authors.lastname: Kraal
mesh_terms: D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology*; D002455:Cell Division / Q000502:physiology; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000166:cytology / Q000187:drug effects / Q000235:genetics*; D005816:Genetic Complementation Test; D010444:Peptide Elongation Factor Tu / Q000235:genetics* / Q000378:metabolism; D011728:Pyridones / Q000494:pharmacology; D012269:Ribosomal Proteins / Q000235:genetics* / Q000378:metabolism; D013307:Streptomycin / Q000378:metabolism*; D013489:Suppression, Genetic
chemical_list: D000900:Anti-Bacterial Agents; D011728:Pyridones; D012269:Ribosomal Proteins; C034212:ribosomal protein S12; D010444:Peptide Elongation Factor Tu; C008536:mocimycin; D013307:Streptomycin