Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.25.1 (deoxyribonuclease)
 1,471 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have constructed a cell-free DNA repair system with UV-irradiated SV40 minichromosomes, as described in the accompanying paper (Sugasawa, K, Masutani, C., and Hanaoka, F. (1993) J. Biol. Chem 268, 9098-9104). In this study, we examined DNA repair synthesis by cell extracts from seven xeroderma pigmentosum (XP) complementation groups, A through G. DNA repair synthesis by XP cell extracts was lower than that with repair-proficient human 293 cell extract and did not increase to the level with the latter on increase in the amount of cell extract or the incubation time. The defects of XP cell extracts were complemented by addition of extracts from cells of different complementation groups, indicating that defective proteins in XP-A through G cells are directly involved in DNA repair. Addition of T4 endonuclease V, which is reported to complement defects of XP cells, stimulated DNA repair synthesis by the 293 cell extract, and also complemented the defects of all XP cell extracts. The XPAC gene product was shown to be involved in DNA repair synthesis using anti-xpac serum and xpac protein produced in Escherichia coli. Anti-xpac serum inhibited DNA repair synthesis by the 293 cell extract and xpac protein reversed the inhibition. Furthermore, xpac protein complemented the defects of extracts of two lines of XP-A cells (XP2OSSV and XP12ROSV) but had no effect on the reactions of extracts from cells of other complementation groups. These findings are consistent with previous results obtained in experiments with cells, indicating that our system is useful for analyzing the mechanisms of DNA excision repair in mammalian cells.
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PMID:Cell-free repair of UV-damaged simian virus 40 chromosomes in human cell extracts. II. Defective DNA repair synthesis by xeroderma pigmentosum cell extracts. 838 77

We constructed a double-stranded plasmid containing a single cis, syn-cyclobutane thymine dimer (T[c,s]T) 385 base pairs from the center of the SV40 origin of replication. This circular DNA was replicated in vitro by extracts from several types of human cells. The dimer was placed on the leading strand template of the first replication fork to encounter the lesion. Two-dimensional gel electrophoresis of replication intermediates documented the transient arrest of the replication fork by the dimer. Movement of the replication fork beyond the dimer was recognized by the appearance of a single fork arc in DNA sequences located between the T[c,s]T and the half-way point around the circular template (180 degrees from the origin). Upon completion of plasmid replication, the T[c,s]T was detected by T4 endonuclease V in about one-half (46 +/- 9%) of the closed circular daughter molecules. Our results demonstrate that extracts prepared from HeLa cells and SV40-transformed human fibroblasts (SV80, IDH4), including a cell line defective in nucleotide-excision repair (XPA), were competent for leading strand DNA synthesis opposite the pyrimidine dimer and replication fork bypass. In contrast, dimer bypass was severely impaired in otherwise replication-competent extracts from two different xeroderma pigmentosum variant cell lines.
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PMID:Replication fork bypass of a pyrimidine dimer blocking leading strand DNA synthesis. 915 57

Genetic information is frequently disturbed by introduction of modified or mismatch bases into duplex DNA, and hence all organisms contain DNA repair systems to restore normal genetic information by removing such damaged bases or nucleotides and replacing them by correct ones. The understanding of this repair mechanism is a central subject in cell biology. This review focuses on the three-dimensional structural views of damaged DNA recognition by three proteins. The first protein is T4 endonuclease V (T4 endo V), which catalyzes the first reaction step of the excision repair pathway to remove pyrimidine-dimers (PD) produced within duplex DNA by UV irradiation. The crystal structure of this enzyme complexed with DNA containing a thymidine-dimer provided the first direct view of DNA lesion recognition by a repair enzyme, indicating that the DNA kink coupled with base flipping-out is important for damaged DNA recognition. The second is very short patch repair (Vsr) endonuclease, which recognizes a TG mismatch within the five base pair consensus sequence. The crystal structure of this enzyme in complex with duplex DNA containing a TG mismatch revealed a novel mismatch base pair recognition scheme, where three aromatic residues intercalate from the major groove into the DNA to strikingly deform the base pair stacking but the base flipping-out does not occur. The third is human nucleotide excision repair (NER) factor XPA, which is a major component of a large protein complex. This protein has been shown to bind preferentially to UV- or chemical carcinogen-damaged DNA. The solution structure of the XPA central domain, essential for the interaction of damaged DNA, was determined by NMR. This domain was found to be divided mainly into a (Cys)4-type zinc-finger motif subdomain for replication protein A (RPA) recognition and the carboxyl terminal subdomain responsible for DNA binding.
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PMID:Three-dimensional structural views of damaged-DNA recognition: T4 endonuclease V, E. coli Vsr protein, and human nucleotide excision repair factor XPA. 1094 33

The ability to detect the most common type of UV-induced mutation, the C to T transition, at the previously characterized hotspot at position 99 of the supF gene has been demonstrated in a selectively irradiated reversion vector, pLS189(Rev). The supF region was amplified, irradiated with 500J/m(2) UVC or unirradiated, and ligated into the pLS189(Rev) plasmid. A portion of ligated product plasmid containing the irradiated fragment was sensitive to nicking by T4 endonuclease V, indicating the presence of the most common type of UV-induced damage, the pyrimidine dimer. Plasmid containing the irradiated or unirradiated supF gene was replicated completely in cellular extracts from either HeLa or XP-A cells in vitro. Plasmid containing the irradiated supF gene showed an inhibition of total replication to a level similar to those of previous studies with plasmid molecules exposed in their entirety to 40J/m(2). Replication of selectively irradiated plasmid resulted in an average reversion frequency of 0.071% in the two extracts; a 42-fold increase over the average spontaneous reversion frequency of unirradiated plasmid. The reversion frequencies were not significantly different between extracts prepared from HeLa and XP-A cells, indicating that neither the repair status of the cell lines nor the XPA protein itself affect the frequency of C to T transitions at position 99 of the supF gene in plasmid replicated in vitro. These data indicate that the plasmid pSL189(Rev), containing the selectively UV-irradiated supF gene, is a useful and sensitive tool to study mutagenesis at a specific site. This approach may be applicable to the investigation of other environmental DNA-damaging agents, by allowing the target gene to be selectively damaged while maintaining the ability of the plasmid to replicate completely. Such a system, amenable to biochemical manipulation, may be very valuable in elucidating the function of novel proteins in the process of mutagenesis.
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PMID:In vitro replication and mutagenesis of a novel reversion vector with selective DNA damage in the supF gene. 1133 80