Well-ordered three-dimensional crystals of 70 S ribosomes and 30 S ribosomal subunits from extremely thermophilic bacteria Thermus thermophilus have been obtained. Positively stained thin sections of the crystals have been analyzed by electron microscopy. Redissolved crystalline ribosomes and small ribosomal subunits reveal sedimentation constants of 70 S and 30 S, respectively, and are functionally active in the poly(U)-system.
The unusually high RNA-binding affinity of MjaL1 might be explained by the exposure of its highly conserved regions. The open conformation of MjaL1 is strongly stabilized by nonconserved interdomain interactions and suggests that the closed conformations of L1 (as in TthL1) open upon RNA binding. Comparison of the two L1 protein structures reveals a high conformational variability of this ribosomal protein. Determination of the MjaL1 structure offers an additional variant for fitting the L1 protein into electron-density maps of the 50S ribosomal subunit.
The gene of the ribosomal protein S8 from Thermus thermophilus VK1 has been isolated from a genomic library by hybridization of an oligonucleotide coding for the N-terminal amino acid sequence of the protein, amplified by PCR and sequenced. Nucleotide sequence reveals an open reading frame coding for a protein of 138 amino acid residues (Mr 15 839). The codon usage shows that 94% of the codons possess G or C in the third position, and agrees with the preferential usage of codons of high G+C content in the bacteria of the genus Thermus. The amino acid sequence of the protein shows 48% identity with the protein from Escherichia coli. Ribosomal protein S8 from 7: thermophilus has been expressed in E. coli under the control of the T7 promoter and purified to homogeneity by heat treatment of the extract followed by cation-exchange chromatography. Conditions were defined in which 7: thermophilus protein S8 binds specifically an homologous 16s rRNA fragment containing the putative S8 binding site with an apparent association constant of 5x10' M-'. The overexpressed protein binds the rRNA with the same affinity as that extracted from 7: thermophilus, indicating that the thermophilic protein is correctly folded in E. coli. The specificity of this binding is dependent on the ionic strength. The protein S8 from 7: thermophilus recognizes the E. coli rRNA binding site as efficiently as the S8 protein from E. coli. This result agrees with sequence comparisons of the S8 binding site on the small subunit rRNA from E. coli and from 7: thermophilus, showing strong similarities in the regions involved in the interaction. It suggests that the structural features responsible for the recognition are conserved in the mesophilic and thermophilic eubacteria, despite structural pecularities in the thermophilic partners conferring thermostability.Although crystals of ribosomes as well as of ribosomal particles diffracting at correct resolution have been obtained (see e.g. Yonath et al., 1990;Yusupov et al., 1991), determination at atomic level of the structure of such multimolecular ribonucleoprotein complexes is obviously a long-term pro-
A new crystalline form of 30 S ribosomal subunits from an extremely thermophilic bacterium Thermus thermophilus has been obtained. Positively stained ultrathin sections of the crystals have been analysed by electron microscopy. The crystals show X-ray diffraction to about 12 A resolution in a synchroton beam at 0°C.Ribosome; 30 S ribosomal subunit; Threedimensional crystal; X-ray analysis; (Thermus fherrnophilus)
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.