Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.1.99.3 (PRE)
 1,923 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cells defective in uracil-DNA glycosylase (ung::Tn10) were used in two ways to reveal differences in select point mutations (GC to AT transitions) within the seven-tRNA operon of E. coli. The mutations were indicated as de novo or converted glutamine tRNA suppressor mutations in the genes glnU and/or glnV: the kinetics of photoenzymatic monomerization of pyrimidine dimers quantitated by ung-dependent UV mutagenesis indicated more rapid repair of dimers at sites for converted suppressor mutation than of dimers at sites for de novo suppressor mutation, and spontaneous deamination of cytosine was considerably more frequent at sites for converted suppressor mutation than at sites for de novo suppressor mutation. To explain these results we suggest the physical structure of the DNA in vivo is different at different sites in the seven-tRNA operon. The non-transcribed strand including specifically the anticodon region of the site for converted suppressor mutation may frequently be looped out in a single strand so that a T = C dimer is more accessible to DNA photolyase or a free cytosine residue of non-irradiated DNA is in an aqueous environment conducive to deamination. In addition, we analysed the spontaneous de novo suppressor mutation data to determine an estimate for the in vivo rate of cytosine deamination in double strand DNA of 3.2 X 10(-13)/sec.
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PMID:Mutation probe of gene structure in E. coli: suppressor mutations in the seven-tRNA operon. 353 75

The rate of UV-induced deamination of cytosine to uracil at a specific site in double-stranded (ds) DNA was monitored using a genetic reversion assay. M13mp2C141 ds DNA was exposed to 160 J/m2 UV (254 nm), incubated at 37 degrees C, pH 7.4, for various time intervals to allow for deamination, and treated with Escherichia coli photolyase in the presence of 365 nm light to reverse cyclobutane-type pyrimidine dimers. Upon transfection into uracil-glycosylase deficient (ung-) E. coli cells, the mutation (i.e., reversion) frequencies in the CCCC target sequence increased greatly with post-UV time of incubation at 37 degrees C, nearly doubling every day that the DNA had been held at 37 degrees C. After 8 days, the reversion frequencies had increased by two orders of magnitude upon transfection into ung- cells, relative to isogenic ung+ cells, indicating that most of the mutations arising in UV/photolyase-treated ds DNA were C-->T mutations mediated by a uracil intermediate. Sequencing of the revertants revealed that all mutations were single C-->T or tandem double CC-->TT mutations. An increasing percentage of tandem double CC-->TT mutations was found with longer post-UV incubation times, yet none occurred if the post-UV delay time step was omitted before photoreversal. After a 4-day delay between UV and photoreversal at 37 degrees C, greater than 84% of the total revertants had tandem double CC-->TT mutations. Thus, the generation of a tandem double mutation is a time-dependent process that arises in DNA after the initial UV exposure. The rate of appearance (with a pseudo-first-order rate constant ca. 10(-6) s-1) of tandem double mutations during incubation of UV-irradiated DNA is inconsistent with two random, independently occurring mutational events and suggests a concerted deamination of both residues in a tandem cytosine pyrimidine (C < > C) dimer. Considering that deamination in a C < > C dimer occurred here with a half-life of ca. 5 days, in contrast to the measured half-life of ca. 20,000 years for spontaneous (non-UV-treated) cytosine deamination for the same target, these studies show that the formation of pyrimidine dimers in DNA increases the rate of deamination by six orders of magnitude, leading to the accelerated formation of single C-->T and tandem double CC-->TT mutations.
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PMID:Accelerated deamination of cytosine residues in UV-induced cyclobutane pyrimidine dimers leads to CC-->TT transitions. 875 82