Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have used in vitro DNA replication systems from human HeLa cells and monkey CV-1 cells to replicate a UV-damaged simian virus 40-based shuttle vector plasmid, pZ189. We found that replication of the plasmid was inhibited in a UV fluence-dependent manner, but even at UV fluences which caused damage to essentially all of the plasmid molecules some molecules became completely replicated. This replication was accompanied by an increase (up to 15-fold) in the frequency of mutations detected in the supF gene of the plasmid. These mutations were predominantly G:C-->A:T transitions similar to those observed in vivo. Treatment of the UV-irradiated plasmid DNA with Escherichia coli photolyase to reverse pyrimidine cyclobutane dimers (the predominant UV-induced photoproduct) before replication prevented the UV-induced inhibition of replication and reduced the frequency of mutations in supF to background levels. Therefore, the presence of pyrimidine cyclobutane dimers in the plasmid template appears to be responsible for both inhibition of replication and mutation induction. Further analysis of the replication of the UV-damaged plasmid revealed that closed circular replication products were sensitive to T4 endonuclease V (a pyrimidine cyclobutane dimer-specific endonuclease) and that this sensitivity was abolished by treatment of the replicated DNA with E. coli photolyase after replication but before T4 endonuclease treatment. These results demonstrate that these closed circular replication products contain pyrimidine cyclobutane dimers. Density labeling experiments revealed that the majority of plasmid DNA synthesized in vitro in the presence of bromodeoxyuridine triphosphate was hybrid density whether or not the plasmid was treated with UV radiation before replication; therefore, replication of UV-damaged templates appears to occur by the normal semiconservative mechanism. All of these data suggest that replication of UV-damaged templates occurs in vitro as it does in vivo and that this replication results in mutation fixation.
Mol Cell Biol 1993 Jan
PMID:Replication and mutagenesis of UV-damaged DNA templates in human and monkey cell extracts. 841 49

Bis(1,10-phenanthroline)-copper(I) ion (OP2Cu+) binds reversibly to B-DNA and makes single-stranded cuts by oxidative attack on the deoxyribose moiety. The deoxyribonuclease activity is sequence-dependent yet not nucleotide-specific at the cutting site. OP2Cu+ sequence specificity was analysed in terms of local variations of DNA stability. Kinetic constants of strand cleavage were measured at sequence positions on the two strands and converted into activation free energies of the cleavage reaction. DNA unwinding free energies were calculated from the base sequence using B-DNA stacking parameters for calculations. The two free-energy variations were statistically compared for a series of DNA restriction fragments bearing the binding sites of regulatory proteins and representing a total of 345 DNA base positions. This study shows that the mean activation free energy of strand cleavage at a pair of opposing sugars across the DNA minor groove varies like the unwinding free energy of the DNA sequence delimited by opposing sugars (3 to 4 bp). A statistical equality between the two free-energy variations is demonstrated when considering the sum of the two cleavage events at the opposing sugars. Systematic deviations between the two free-energy distributions were observed at specific sequences, including polypurine-polypyrimidine tracts (AnTm/AmTn, CnTmCp/GpAmGn), alternating purine-pyrimidine tracts ((TA)n/(TA)n, (TG)n/(CA)n) and at certain G+C-rich triplets (GGC, GCC and CGC). The physical significance of these observations is discussed and a model of OP2Cu+ binding and cleavage specificity based on the free-energy equality is proposed.
J Mol Biol 1996 Jul 26
PMID:DNA-stacking interactions determine the sequence specificity of the deoxyribonuclease activity of 1,10-phenanthroline-copper ion. 875 18

The ATP-dependent deoxyribonuclease enzyme complex (AddAB) of Bacillus subtilis possesses two consensus ATP-binding sequences, located in the N-terminal region of both subunits. The highly conserved lysine residues in both consensus ATP-binding sequences were replaced by glycine, resulting in the mutant enzyme complexes AddAB-A-K36G (AddA*B) and AddAB-B-K14G (AddAB*). The mutation in subunit AddA reduced DNA repair and chromosomal transformation, and abolished bacteriophage PBS1-mediated transduction. This mutation also resulted in a complete loss of the ATP-dependent exonuclease and helicase activity. In contrast, the mutation in subunit AddB had only marginal effects. The recF and addAB genes are not required for transformation with plasmid DNA, but have overlapping activities in transformation with chromosomal DNA. By contrast to RecF, the AddAB enzyme is essential for PBS1-mediated transduction. However, recF has a more important function with respect to DNA repair than addAB.
Mol Microbiol 1996 Sep
PMID:Replacement of the lysine residue in the consensus ATP-binding sequence of the AddA subunit of AddAB drastically affects chromosomal recombination in transformation and transduction of Bacillus subtilis. 888 69

A novel enzymatic activity on nucleic acids was discovered in both muscle type (MT) and erythrocyte or common type (CT) isoforms of acylphosphatase, an enzyme that was previously known as a hydrolase (E.C.3.6.1.7). Both deoxyribonucleic and ribonucleic hydrolitic activity were assayed on a variety of substrates. Our results demonstrate that acylphosphatase possesses both Mg++ dependent deoxyribonuclease and ribonuclease activities, at pH ranging from 5.0 to 6.8. Furthermore, we present evidences, for both isoenzymatic forms, of the coexistence of exonucleolytic and endonucleolytic activities on DNA.
Biochem Mol Biol Int 1996 Sep
PMID:Characterization of a novel nucleolytic activity of acylphosphatases. 888 72

We have established a stably transformed human neuroblastoma cell line (MC65) that conditionally expresses a C-terminal derivative of the amyloid beta protein precursor (beta PP) termed S beta C (a fusion protein composed of the amino-17 and carboxyl-99 residues of beta PP). Conditional expression of S beta C (mediated by the withdrawal of tetracycline from the culture medium) induces pronounced nuclear DNA fragmentation and cytotoxicity in this cell line. These effects are enhanced by hyperoxygen and suppressed by hypooxygen and antioxidants. This cell line is relatively insensitive to the extracellular application of amyloid beta 25-35, and coculture experiments suggest that this cytotoxicity is mediated by an intracellular process. These findings suggest that the overexpression of the C-terminal domain of beta PP can disrupt normal cellular processes in these cells in such a way as to induce a directed (deoxyribonuclease-mediated) mechanism of cell death. This process appears to be modulated and/or mediated by a reactive oxygen specie(s) (ROS). Consistent with a role for ROS in the process of S beta C-mediated toxicity, we have found that the MC65 cell line is hypersensitive to oxidative stress and that it is this sensitivity that appears (at least in part) to underlie its susceptibility to S beta C.
Mol Chem Neuropathol
PMID:Neurodegenerative mechanisms in Alzheimer disease. A role for oxidative damage in amyloid beta protein precursor-mediated cell death. 897 93

To evaluate the specificity and applicability to the study of human tumor cells of the reverse transcription (RT) in situ PCR and RT polymerase chain reaction (PCR) in situ hybridization techniques, we examined five melanoma cell lines and five nonmelanoma lines for tyrosinase mRNA using primers specific for tyrosinase. Each procedural step was optimized and minutely controlled, and results from the in situ techniques and solution-phase RT-PCR were compared. All melanoma lines showed a specific pattern of perinuclear cytoplasmic reaction not seen in nonmelanoma lines. There was exact agreement between the results from the RT in situ PCR and RT-PCR in situ hybridization techniques and those from solution-phase RT-PCR. Ribonuclease digestion abolished cytoplasmic staining, as did omission of the reverse transcriptase step. Nuclear staining was seen in melanoma and nonmelanoma lines, apparently as a result of DNA synthesis from repair-replication and mispriming or nonspecific amplification. Neither high concentrations of deoxyribonuclease nor long incubation periods abolished this effect completely. Demonstration of cytoplasmic mRNA by RT in situ PCR and RT-PCR in situ hybridization specifically identifies cells of melanocytic lineage.
Diagn Mol Pathol 1997 Feb
PMID:Demonstration of cytoplasmic tyrosinase mRNA in tissue-cultured cells by reverse transcription (RT) in situ polymerase chain reaction (PCR) and RT PCR in situ hybridization. 902 34

The rat androgen-binding protein (ABP) gene is transcriptionally regulated from two promoters: the P1 promoter regulates expression of transcripts starting at exon 1, whereas P(A) regulates transcripts containing exon A. The P1 promoter directs cell-specific gene regulation of ABP secreted by Sertoli cells. In this study, the Sertoli cell-regulatory sequences of P1 were further examined using a luciferase reporter system with three cell lines, including a Sertoli cell line (MSC-1) that expresses the ABP gene. Deletion mapping experiments determined that the sequences required for full activity in MSC-1 cells were included within 619 bp of the start site and identified several regions that demonstrated increased luciferase activity: the -583 bp to -564 bp, -503 bp to -484 bp, and -114 bp to -65 regions. The activities contributed by each region were much higher (up to 120-fold) in MSC-1 cells than in MA10 Leydig or NIH3T3 fibroblast cells. Nuclear-binding proteins and their binding sequences were identified using several molecular biology techniques. Complexes formed by nuclear proteins of MSC-1, MA10, and NIH3T3 cells, which bind specifically to the -114 to -65-bp region, were identified using gel retardation assays. Furthermore, the inverted repeat sequence in this region, 5'-AGGGTCAGTGTCCCT-3' was identified by deoxyribonuclease (DNase) I footprinting. The regulatory element contained within the -503 to -484-bp region was identified by scanning mutagenesis, but no protein was found that bound to this sequence by gel retardation or DNase I protection assays. This element is characterized by the core sequence, 5'-GGAGGC-3'. The third regulatory region (residues -583 to -564) bound a protein complex that retarded mobility of the free DNA probe in a gel shift assay. Using several techniques, the binding sequence was identified as 5'-TTCATAGTATCCATTAAAC-3'. In summary, these data have identified several transcriptional regulatory sequences and their binding proteins, which appear to play a role in the Sertoli cell-specific expression of the ABP gene.
Mol Endocrinol 1997 Aug
PMID:DNA sequences and their binding proteins required for Sertoli cell-specific transcription of the rat androgen-binding protein gene. 925 28

To understand the causes of CAG repeat tract changes that occur in the passage of human disease alleles, we are studying the effect of replication and repair mutations on CAG repeat tracts embedded in a yeast chromosome. In this report, we examine the effect of a mutation in the RTH1/RAD27 gene encoding a deoxyribonuclease needed for removal of excess nucleotides at the 5'-end of Okazaki fragments. Deletion of the RTH1/RAD27 gene has two effects on CAG tracts. First, the rth1/rad27 mutation destabilizes CAG tracts. Second, although most tract length changes in wild-type yeast cells are tract contractions, approximately half of the changes that occur as a result of the rth1/rad27 mutation are expansions of one or more repeat units. These results support the hypothesis that tract expansions that occur during passage of human disease alleles bearing expanded CAG tracts result from excess DNA synthesis on the lagging strand of replication.
Hum Mol Genet 1998 Jan
PMID:Expansions of CAG repeat tracts are frequent in a yeast mutant defective in Okazaki fragment maturation. 938 5

One of the major DNA repair pathways is base excision repair, in which DNA bases that have been damaged by endogenous or exogenous agents are removed by the action of a class of enzymes known as DNA glycosylases. One subset of the known DNA glycosylases has an associated abasic lyase activity that generates a phosphodiester bond scission. The base excision pathway is completed by the sequential action of abasic endonucleases, DNA polymerases, and DNA ligases. Base excision repair of ultraviolet (UV) light-induced dipyrimidine photoproducts has been described in a variety of prokaryotic and eukaryotic organisms and phages. These enzymes vary significantly in their exact substrate specificity and in the catalytic mechanism by which repair is initiated. The prototype enzyme within this class of UV-specific DNA glycosylases is T4 endonuclease V. Endonuclease V holds the distinction of being the first glycosylase (1) to have its structure solved by X-ray diffraction of the enzyme alone as well as in complex with pyrimidine dimer-containing DNA, (2) to have its key catalytic active site residues identified, and (3) to have its mechanism of target DNA site location determined and the biological relevance of this process established. Thus, the study of endonuclease V has been critical in gaining a better understanding of the mechanisms of all DNA glycosylases.
Prog Nucleic Acid Res Mol Biol 1999
PMID:The initiation of DNA base excision repair of dipyrimidine photoproducts. 993 54

This report demonstrates that Gadd45, a p53-responsive stress protein, can facilitate topoisomerase relaxing and cleavage activity in the presence of core histones. A correlation between reduced expression of Gadd45 and increased resistance to topoisomerase I and topoisomerase II inhibitors in a variety of human cell lines was also found. Gadd45 could potentially mediate this effect by destabilizing histone-DNA interactions since it was found to interact directly with the four core histones. To evaluate this possibility, we investigated the effect of Gadd45 on preassembled mononucleosomes. Our data indicate that Gadd45 directly associates with mononucleosomes that have been altered by histone acetylation or UV radiation. This interaction resulted in increased DNase I accessibility on hyperacetylated mononucleosomes and substantial reduction of T4 endonuclease V accessibility to cyclobutane pyrimidine dimers on UV-irradiated mononucleosomes but not on naked DNA. Both histone acetylation and UV radiation are thought to destabilize the nucleosomal structure. Hence, these results imply that Gadd45 can recognize an altered chromatin state and modulate DNA accessibility to cellular proteins.
Mol Cell Biol 1999 Mar
PMID:Gadd45, a p53-responsive stress protein, modifies DNA accessibility on damaged chromatin. 1002 55


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