In vitro transcribed thymidylate synthase (TS) mRNA which is 100% substituted with 5-fluorouracil (FUra) was analyzed for changes in mRNA secondary structure, for alterations in translational efficiency, and for evidence of translational miscoding in vitro. FUra substitution in TS mRNA results in an altered migration pattern in non-denaturing RNA gels and in decreased hyperchromicity in RNA mel...

8-(hydroxymethyl)-3,N-etheno-dC (8-HM-εC), is a mutagen and animal carcinogen resulting from the reaction of dC with glycidaldehyde. It was synthesized and its phosphoramidite was incorporated site-specifically into a defined 25-mer oligonucleotide. In this study the mutagenic potential of this compound was investigated in an in vitro primer-template extension assay using four mammalian DNA pol...

Negamycin is a natural product with broad-spectrum antibacterial activity and efficacy in animal models of infection. Although its precise mechanism of action has yet to be delineated, negamycin inhibits cellular protein synthesis and causes cell death. Here, we show that single point mutations within 16S rRNA that confer resistance to negamycin are in close proximity of the tetracycline bindin...

A site-specifically modified oligonucleotide containing a single 2'-deoxyribonolactone lesion was used as a template for primer extension reactions catalyzed by M-MuLV reverse transcriptase (RT) and by the Klenow fragments of Escherichia coli DNA polymerase proficient (KF exo(+)) or deficient (KF exo(-)) in exonuclease activity. Analysis of the extension products in the presence of the four dNT...

Deamination of nucleobases in DNA and RNA results in the formation of xanthine (X), hypoxanthine (I), oxanine, and uracil, all of which are miscoding and mutagenic in DNA and can interfere with RNA editing and function. Among many forms of nucleic acid damage, deamination arises from several unrelated mechanisms, including hydrolysis, nitrosative chemistry, and deaminase enzymes. Here we presen...

Aminoglycosides are chemically diverse, broad-spectrum antibiotics that target functional centers within the bacterial ribosome to impact all four principle stages (initiation, elongation, termination, and recycling) of the translation mechanism. The propensity of aminoglycosides to induce miscoding errors that suppress the termination of protein synthesis supports their potential as therapeuti...

1,N(6)-ethanoadenine (EA) forms through the reaction of adenine in DNA with the antitumor agent 1,3-bis(2-chloroethyl)-1-nitrosourea, a chemotherapeutic used to combat various brain, head, and neck tumors. Previous studies of the toxic and mutagenic properties of the DNA adduct EA have been limited to in vitro experiments using mammalian polymerases and have revealed the lesion to be both misco...

The carcinogenic and mutagenic effects of the aromatic amines are believed to depend on their covalent modification of DNA, primarily through the formation of adducts at C8 of guanine. The actual biologic and biochemical responses to these adducts can be envisioned as the consequence of the abilities of the cell to repair the lesions, with or without fidelity, and the introduction of errors thr...

The temporal regulation of DNA repair during synchronous cell proliferation was examined in normal human skin fibroblasts and in Bloom's syndrome skin fibroblasts. Normal human cells regulated DNA repair in a defined temporal sequence prior to the induction of DNA replication. Nucleotide-excision repair was stimulated prior to the induction of base-excision repair, which itself was increased pr...

trans-Translation is an unusual translation in which tmRNA plays a dual function as a tRNA and an mRNA to relieve the stalled translation on the ribosome. In this study, we examined the effects of an aminoglycoside antibiotic, paromomycin, on several tmRNA-related events in vitro. The results of a chemical footprinting study indicated that paromomycin molecules bind tmRNA at G332/G333 in the tR...