Streptomycin-resistant and streptomycin-dependent mutants of the extreme thermophile Thermus thermophilus

We have isolated spontaneous streptomycin-resistant, streptomycin-dependent and streptomycin-pseudo-dependent mutants of the thermophilic bacterium Thermus thermophilus IB-21. All mutant phenotypes were found to result from single amino acid substitutions located in the rpsL gene encoding ribosomal protein S12. Spontaneous suppressors of streptomycin dependence were also readily isolated. Thermus rpsL, mutations were found to be very similar to rpsL mutations identified in mesophilic organisms. This similarity affords greater confidence in the utility of the crystal structures of Thermus ribosomes to interpret biochemical and genetic data obtained with Escherichia coli ribosomes. In the X-ray crystal structure of the T. thermophilus HB8 30 S subunit, the mutated residues are located in close proximity to one another and to helices 18, 27 and 44 of 16 S rRNA. X-ray crystallographic analysis of ribosomes from streptomycin-resistant, streptomycin-pseudo-dependent and streptomycin-dependent mutants described here is expected to reveal fundamental insights into the mechanism of tRNA selection, translocation, and conformational dynamics of the ribosome. (C) 2001 Academic Press.