Functional interactions within 23S rRNA involving the peptidyltransferase center.
A molecular genetic approach has been employed to investigate functional interactions within 23S rRNA. Each of the three base substitutions at guanine 2032 has been made. The 2032A mutation confers resistance to the antibiotics chloramphenicol and clindamycin, which interact with the 23S rRNA peptidyltransferase loop. All three base substitutions at position 2032 produce an erythromycin-hypersensitive phenotype. The 2032 substitutions were compared with and combined with a 12-bp deletion mutation in domain II and point mutations at positions 2057 and 2058 in the peptidyltransferase region of domain V that also confer antibiotic resistance. Both the domain II deletion and the 2057A mutation relieve the hypersensitive effect of the 2032A mutation, producing an erythromycin-resistant phenotype; in addition, the combination of the 2032A and 2057A mutations confers a higher level of chloramphenicol resistance than either mutation alone. 23S rRNAs containing mutations at position 2058 that confer clindamycin and erythromycin resistance become deleterious to cell growth when combined with the 2032A mutation and, additionally, confer hypersensitivity to erythromycin and sensitivity to clindamycin and chloramphenicol. Introduction of the domain II deletion into these double-mutation constructs gives rise to erythromycin resistance. The results are interpreted as indicating that position 2032 interacts with the peptidyltransferase loop and that there is a functional connection between domains II and V.
Douthwaite
S
S
Department of Molecular Biology, Odense University, Denmark.
D010582:Bacteriophage lambda / Q000235:genetics; D001482:Base Composition; D001483:Base Sequence; D002702:Chloramphenicol Resistance; D002981:Clindamycin / Q000494:pharmacology; D004252:DNA Mutational Analysis; D004352:Drug Resistance, Microbial / Q000235:genetics*; D004917:Erythromycin / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D010458:Peptidyl Transferases / Q000378:metabolism*; D010957:Plasmids / Q000187:drug effects / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012338:RNA, Ribosomal, 23S / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism
D012338:RNA, Ribosomal, 23S; D002981:Clindamycin; D004917:Erythromycin; D010458:Peptidyl Transferases