Effects of mutagenesis of C912 in the streptomycin binding region of Escherichia coli 16S ribosomal RNA
| Autor: | Elizabeth A. De Stasio, Anne L. Frattali, Albert E. Dahĺberg, Michael K. Flynn |
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| Rok vydání: | 1990 |
| Předmět: |
Models
Molecular Molecular Sequence Data Mutant Biophysics Mutagenesis (molecular biology technique) Biology medicine.disease_cause Biochemistry Microbiology Plasmid Structural Biology Ribosomal protein RNA Ribosomal 16S Escherichia coli Genetics medicine Protein biosynthesis Binding Sites Base Sequence Drug Resistance Microbial Ribosomal RNA Molecular biology Streptomycin Mutagenesis Site-Directed Nucleic Acid Conformation Chromosome Deletion Plasmids medicine.drug |
| Zdroj: | Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression. 1050:27-33 |
| ISSN: | 0167-4781 |
| DOI: | 10.1016/0167-4781(90)90136-p |
| Popis: | Four different mutations were produced at position 912 of Escherichia coli 16S rRNA in the multicopy plasmid pKK3535. Cells transformed with the mutant plasmids were assayed for growth in steptomycin. The U912 mutant conferred low level streptomycin resistance as reported originally by Montandon and co-workers (EMBO J 1986; 5:3705-3708). The G912 mutant also gave low level resistance but, unlike U912, caused significant retardation in growth rate and tended to select for fast-growing revertants. The A912 mutant was without effect on growth rate or streptomycin sensitivity, while deletion of C912 was lethal. Cells with U912 were selected for increased streptomycin resistance (MIC up to 160 micrograms/ml) and then cured of the plasmid. The cured cells retained a higher level of streptomycin resistance (MIC: 80 micrograms/ml) than the original wild type strain (MIC: 10 micrograms/ml), but sequencing by reverse transcriptase showed no evidence of U912 in the cellular 16S rRNA. Thus, recombination of the plasmid-coded U912 mutation into host rrn operons was not the mechanism by which increased streptomycin resistance occurred. The plasmid with U912 was transformed into three different streptomycin-dependent strains to determine whether the rRNA mutation, which presumably alters streptomycin binding, was compatible with S12 mutations which require bound streptomycin in order to function properly. In one strain, no transformants could be isolated, indicating that the plasmid was lethal. The two other streptomycin-dependent strains were transformed, but ribosomes containing the mutant rRNA were non-functional. |
| Databáze: | OpenAIRE |
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