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Summary Spectinomycin resistant (spc r) mutants were obtained by treating the cells of E. coli K12, W3637 with nitrosoguanidine. The compositions of ribosomal proteins were analyzed for six out of eleven such spc r-mutants with chromatography on a carboxymethyl cellulose (CMC) column. The 30s ribosomal subunit from all of the spc r-mutants was found to contain the altered 30-4 protein component, while no difference was detected in 50s ribosomal proteins between spc r and spc s bacteria.Abbreviations used CMC carboxymethyl cellulose - str streptomycin - spc spectinomycin  相似文献   

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Two amber mutations have been mapped inside the spcA-strA region (now called rpsE-rpsL) on the bacterial genome. Derivatives of the transducing phage lambda fus3 carrying each mutation were constructed and assayed in ultraviolet-irradiated bacteria to identify the mutated genes and measure the polarity of the mutations. The data indicated that both mutations, 3162(Am) and 3161(Am), affect genes coding for ribosomal proteins: rplC (L3) and rpsN (S14), respectively. It was shown also that each mutation exerts, inside of its respective operon (S10 and spc units), a relatively strong polar effect on genes distal to the mutated locus.  相似文献   

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Oligonucleotide-directed mutagenesis was used to introduce an A to C transversion at position 523 in the 16S ribosomal RNA gene of Escherichia coli rrnB operon cloned in plasmid pKK3535. E. coli cells transformed with the mutated plasmid were resistant to streptomycin. The mutated ribosomes isolated from these cells were not stimulated by streptomycin to misread the message in a poly(U)-directed assay. They were also restrictive to the stimulation of misreading by other error-promoting related aminoglycoside antibiotics such as neomycin, kanamycin or gentamicin, which do not compete for the streptomycin binding site. The 530 loop where the mutation in the 16S rRNA is located has been mapped at the external surface of the 30S subunit, and is therefore distal from the streptomycin binding site at the subunit interface. Our results support the conclusion that the mutation at position 523 in the 16S rRNA does not interfere with the binding of streptomycin, but prevents the drug from inducing conformational changes in the 530 loop which account for its miscoding effect. Since this effect primarily results from a perturbation of the translational proofreading control, our results also provide evidence that the 530 loop of the 16S rRNA is involved in this accuracy control.  相似文献   

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Cross-linking of streptomycin to the 16S ribosomal RNA of Escherichia coli   总被引:6,自引:0,他引:6  
[3H]Dihydrostreptomycin was cross-linked to the 30S ribosomal subunit from Escherichia coli with the bifunctional reagent nitrogen mustard. The cross-linking primarily involved the 16S RNA. To localize the site of cross-linking of streptomycin to the 16S RNA, we hybridized RNA labeled with streptomycin to restriction fragments of the 16S RNA gene. Labeled RNA hybridized to DNA fragments corresponding to bases 892-917 and bases 1394-1415. These two segments of the ribosomal RNA must be juxtaposed in the ribosome, since there is a single binding site for streptomycin. This region has been implicated both in the decoding site and in the binding of initiation factor IF-3, indicating its functional importance.  相似文献   

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Summary Spontaneous mutants of the petite-negative yeast Kluyveromyces lactis, resistant to the antibiotics chloramphenicol and oligomycin, were isolated and genetically characterized.Three chloramphenicol-resistant mutants showed non-Mendelian inheritance when crossed to sensitive parents.Of 5 oligomycin-resistant strains studied, three exhibited resistance due to the presence of an extrachromosomal mutation. The resistance of the other two deriving from a nuclear and recessive mutation.When two factor crosses in trans configuration were performed between two of the chloramphenicol and the five oligomycin-resistant mutants a polarity in recombination was observed with a predominance of sensitive (OSCS) over resistant (ORCR) reciprocal recombinants.Allelism tests carried out among the oligomycin-resistant mutants indicated the presence of at least two distinct extrachromosomal regions responsible for the resistance.  相似文献   

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Summary The Escherichia coli K12 strain derivative MX 109, carries the strA938 mutant allele. It was isolated as a spontaneous high-level streptomycin resistant mutant, having the additional characteristic of growing slowly at a wide range of temperatures. Thus far, these two phenotypic characteristics have not been genetically segregated and, probably, are the result of a single mutational event at the strA locus.A number of pleiotropic alterations are also observed in cells of MX109 when compared two wild type cells, very likely as a consequence of the expression of the strA938 mutant allele. These are, i) decrease in their polysome content, ii) increase in their RNA to protein ratio, iii) increase in the density of the cells, iv) accumulation during growth of 30S and 50S ribosomal precursors, and v) low efficiency of their ribosomes to support polypeptide synthesis in vitro. Most of these effects are observed at 37° C but are more marked if cells or extracts are incubated at 20° C. These observations point to a reduced efficiency for protein synthesis of the ribosomes of MX109, as the primary effect exerted by the 30S-protein product of the strA938 mutant allele.  相似文献   

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Summary Fragments of drif d 18 DNA with different end-points within the set of structural genes of ribosomal proteins L11 (rplK), L1 (rplA), L10 (rplJ) and L12 (rplL) as well as the (rpoB) and (rpoC) subunits of RNA polymerase have been cloned on plasmids. These plasmids were transformed in host cells which were mutant for each of the genes, enabling expression of both wild-type (plasmid-borne) and mutant (chromosomal) genes to be differentiated. On the basis of these results we propose the following genetic structure for the region: rplK and rplA are in one operon; rplL, rpoB and rpoC are in a second. Our data suggest the possibility that rplJ is by itself in an operon situated between the other two.  相似文献   

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A mutant of Escherichia coli has been isolated which exhibits partial suppression of spectinomycin resistance. The site of mutation is in the streptomycin (strA) region and is closely linked to the spcA gene. However, this gene, which we propose to call mod, is phenotypically distinguishable from both the neomycin-kanamycin (nek) and the ribosomal ambiguity gene (ram). The relative gene order is mod spcA strA. In a cell-free protein synthesizing system, altered ribosomes appear to be responsible for the suppression of spectinomycin resistance caused by mod.  相似文献   

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