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)

Affinity modification of E. coli DNA polymerase I and its Klenow fragment by imidazolides of dNMP (Im-dNMP) and dNTP was studied. DNA polymerase activity of DNA polymerase I was reduced by both Im-dNMP and Im-dNTP. However Im-dNTP does not inactivate of the Klenow fragment. The level of covalent labelling of both enzymes by radioactive Im-dNTP did not exceed 0.01 mol of reagent per mol of enzyme. But the deep inactivation of DNA polymerase I by Im-dNTP was observed. It is likely that this inactivation is due to the formation of intramolecular ether followed by phosphorylation of the carboxyl group. This assumption is strongly supported by the increase of the isoelectrical point of DNA polymerase I after its incubation with Im-dNTP in conditions of enzyme inactivation. All data permit us to suggest that the affinity modification of both enzymes by Im-dNMP and covalent labeling by Im-dNTP takes place without complementary binding of dNTP moiety with the template. However inactivation of DNA polymerase I by Im-dNTP occurs only if the dNTP-moiety is complementary to the template in the template.primer complex. It was shown that His residue was phosphorylated by Im-dNMP and Tyr or Ser residues between Met-802 and Met-848 were phosphorylated by Im-dNTP. We suppose that there are two states of DNA polymerase active site for the binding of dNTPs. One of them is independent on the template, in the other state the dNTP hydrogen bond with the template is formed.
Mol Biol (Mosk)
PMID:[Affinity modification of DNA polymerase I from Escherichia coli and its Klenow fragment with nucleotide imidazolides]. 188 93

Using electroporation with the phage PRD1 genome, we set up a high-frequency DNA transfer system for a linear dsDNA molecule with 5'-covalently linked terminal proteins. The transfer was saturated when more than 100 ng of PRD1 genome was used. Electroporation efficiency was about four orders of magnitude higher than that obtained with transfection. Removal of the terminal protein abolished plaque formation, which could not be rescued by supplying the terminal protein or phage DNA polymerase or both in trans.
Mol Gen Genet 1991 Aug
PMID:High-frequency transfer of linear DNA containing 5'-covalently linked terminal proteins: electroporation of bacteriophage PRD1 genome into Escherichia coli. 188 19

A high molecular weight mitochondrial DNA (mtDNA) replication complex, associated with the mitochondrial membrane, was isolated by sucrose gradient centrifugation from purified wheat embryo mitochondria. This complex comprised the mtDNA as well as enzyme activities involved in the replication and transcription of the organelle genome, such as DNA polymerase, RNA polymerase and topoisomerase type I. The isolated complex is active in mtDNA and mtRNA synthesis in vitro. Electron microscopy and lipid analysis confirmed the membrane origin of this complex. Enzyme activities are resistant to physiological ionic strengths, 0.1-0.2 M KC1, while the membrane-mtDNA association is resistant up to 1 M KC1. DNase treatment of the complex released the DNA polymerase activity while protease treatment solubilized mtDNA, suggesting the direct interaction of mtDNA with membrane protein(s). The use of a novel approach to detect mtDNA fragments specifically retained by the mitochondrial membranes after Sal I digestion of the complex suggests that specific mtDNA sequences anchor mtDNA to mitochondrial membranes.
Plant Mol Biol 1991 Feb
PMID:Isolation from wheat mitochondria of a membrane-associated high molecular weight complex involved in DNA synthesis. 189 1

One variant, aphhs-3 was previously isolated based on a hypersensitivity to nontoxic concentrations of aphidicolin, a specific inhibitor of DNA polymerases-alpha and delta. This variant was found to be more sensitive to temperatures above 35 degrees C and to 10 microM of 3'-azido-3'-deoxythymidine (zidovudine, azidothymidine, or AZT) than the parental 743x cells. DNA polymerase activities in the cell extract or in the partially purified fraction by DEAE-cellulose (DE52) anion exchange column from aphhs-3 were active at 40 degrees C. No significant differences in deoxynucleoside triphosphate pools were observed at 34 degrees C for both the parental 743x and aphhs-3 cells. Revertants were isolated at 39 degrees C: six revertants (aphhs-3-tr1 through aphhs-3-tr6) were obtained without aphidicolin; one revertant aphhs-3-tar (the tar clone) was selected in aphidicolin (0.12 microM). The hypersensitivity to aphidicolin (Aphhs) and AZT (AZThs) was cosegregated in the revertant aphhs-3-tr5 (the tr5 clone), while the tar clone was not AZThs. There was a similar increase in the specific activity of 3H-labeled DNA in all cell lines after additions of [3H]AZT or [3H]thymidine. Additions of purine or pyrimidine arabinosides (araT, araC, and araA) to all cell lines resulted in a similar cytotoxicity, suggesting the anabolism of dTTP was not defective in the tr5 clone. The spontaneous mutation rate at the hypoxanthine-guanine phosphoryltransferase locus using replating techniques and 6-thioguanine resistance selection was less than or equal to 5 x 10(-7), 2.2 x 10(-6), or 1.3 x 10(-6) per generation for the tr5, 743x, or tar cell lines, respectively. Most importantly, DNA polymerase activities in the cell extract of the revertant tr5 clone were inhibited by 0.5 microM AZTTP. In contrast, no inhibition was observed in those of the parental 743x and revertant tar cells. The cosegregation of both Aphhs and AZThs in the tr5 revertant suggests that these two phenotypes may be a result of the same mutational event.
Somat Cell Mol Genet 1991 Jan
PMID:Mutants from V79 fibroblasts exhibiting hypersensitivity to aphidicolin and 3'-azido-3'-deoxythymidine. 190 Jan 32

DNA polymerases from the malaria parasite Plasmodium berghei were purified more than 50-fold. Several distinct enzymatic activities were isolated that could be distinguished by the use of various specific DNA polymerase inhibitors. In particular, subdivision into an aphidicolin-sensitive and an aphidicolin-resistant group was possible. Further analysis allowed a better comparison with host DNA polymerases and indicated that one aphidicolin-sensitive DNA polymerase resembled DNA polymerase alpha displaying processive DNA synthesis and using RNA primers, whereas another aphidicolin-sensitive DNA polymerase was distributive and only used DNA primers. Marked differences from the host enzymes do exist, however, such as insensitivity to BuPdGTP. Another P. berghei DNA polymerase was isolated that showed characteristics of a DNA polymerase beta-like enzyme, but which differed from host DNA polymerase beta in its insensitivity to dideoxynucleotides.
Mol Biochem Parasitol 1991 Apr
PMID:Purification and characterization of DNA polymerases from Plasmodium berghei. 190 44

In Saccharomyces cerevisiae, three different DNA polymerase complexes, POLI, POLII and POLIII, are known to be involved in DNA replication. The catalytic subunit of POLIII is encoded by the essential CDC2 gene. The existence of different thermosensitive noncomplementing mutants of CDC2 offers the possibility of using a genetic approach to investigate the involvement of POLIII in induced gene conversion. When cdc2 heteroallelic cells were irradiated and incubated under restrictive conditions, almost no induction of thermoresistant cells could be detected, suggesting an essential role for POLIII in mitotic gene conversion events.
Mol Gen Genet 1991 Oct
PMID:Possible involvement of the yeast POLIII DNA polymerase in induced gene conversion. 194 22

The Schizosaccharomyces pombe POL3 gene was isolated by sequence homology with a region of the Saccharomyces cerevisiae POL3 gene, the only gene sequenced to date encoding the catalytic subunit of eukaryotic DNA polymerase delta. The fission yeast POL3 gene contains a 52 base-pair (bp) intron and encodes a 3600 bp transcript the 5'-end of which is located 32 bp upstream from the initiation codon. The polypeptides predicted from budding and fission yeast POL3 genes share 52% of conserved amino acid residues and have a 60% identical central region. This structural conservation of the catalytic subunit of DNA polymerases delta is probably related to functional constraints. A portion of the most conserved region was used to raise antibodies against an S. pombe polymerase delta/beta-galactosidase fusion protein expressed in Escherichia coli. The purified antibodies recognized a 123,000 Da protein in S. pombe wild-type cell extracts and inhibited an aphidicolin-sensitive DNA polymerase activity that was distinct from DNA polymerase alpha. The antibodies also detected a 140,000 Da protein in extracts from different proliferating mammalian cells, indicating that the catalytic subunits of DNA polymerase delta are highly conserved between yeast and higher eukaryotes.
J Mol Biol 1991 Nov 20
PMID:Characterization of the POL3 gene product from Schizosaccharomyces pombe indicates inter-species conservation of the catalytic subunit of DNA polymerase delta. 196 Jul 23

Since 1983, a series of experimental and clinical studies have been carried out on the possibility of enhancing the chemotherapy effectiveness in breast cancer by expanding the fraction of cycling cells. Theoretically estrogens should recruit breast cancer cells and this fact should result in a higher killing efficiency of antiproliferative drugs. Actually it has been clearly shown, by means of the thymidine labeling index and primer-dependent alpha-DNA polymerase assay, that low doses of diethylstilbesterol are able to increase the tumor proliferative activity of human breast cancer in vivo (estrogenic recruitment). Three randomized trials have been carried out (one in locally advanced and two in metastatic breast cancer) comparing conventional polychemotherapy vs chemotherapy with estrogenic recruitment. Only limited advantages have been observed in these trials. Searching for new modalities of kinetic manipulation of tumors, recombinant human growth hormone has been employed in a pilot study: the preliminary results indicate that it largely enhances tumor proliferative activity, suggesting the possibility of employing a growth factor system to increase chemosensitivity.
J Steroid Biochem Mol Biol 1990 Dec 20
PMID:In vivo manipulation of human breast cancer growth by estrogens and growth hormone: kinetic and clinical results. 198 Oct 14

Primer protein (PP) of bacteriophages M2 and phi 29 contains an Arg-Gly-Asp (RGD) sequence. The RGD-mediated protein-protein interaction in protein-primed DNA replication of M2 was studied in vitro using three purified and indispensable components: PP, DNA polymerase (POL) and template DNA linked to terminal protein (TP). PP competed with a synthetic RGD peptide for binding to the template DNA-TP complex (TP-DNA). In addition, POL bound to template TP-DNA only when complexed with PP. These results indicate that the RGD sequence of PP is responsible for the interaction of the PP-POL complex with TP-DNA, which contains the initiation site for the protein priming of DNA synthesis. At the moment when PP converts to TP upon linking the first deoxynucleotide, a conformational change results in exposure of the RGD binding site.
Mol Gen Genet 1991 Apr
PMID:Primer protein of bacteriophage M2 exposes the RGD receptor site upon linking the first deoxynucleotide. 203 31

The study of plant DNA polymerases lags far behind that concerning their animal or yeast counterpart. In this work we describe the first extensive purification to apparent homogeneity, as well as a detailed biochemical and immunological characterization, of a low molecular weight DNA polymerase (DNA polymerase CI) purified from wheat embryos. The monomeric enzyme is a basic protein having a molecular weight of 52 kDa. Polyclonal antibodies raised in rabbits against DNA polymerase CI did not inhibit animal DNA polymerases alpha and beta or wheat DNA polymerase A, whereas wheat DNA polymerases CII and B were much less affected than the CI enzyme. Several properties of enzyme CI were studied. Some known inhibitors of DNA polymerase activity including aphidicolin, phosphonoacetic acid and heparin, did not affect DNA polymerase CI while the activity of this enzyme was strongly inhibited by ddTTP and N-ethylmaleimide. The polyamine spermine decreased markedly the enzyme activity, while spermidine produced a strong stimulation at the same concentrations that spermine inhibited the enzyme. The best template for this enzyme is poly dA-oligo dT, although polymerase CI can recognize significantly some synthetic polyribonucleotide templates (poly rC-oligo dG, poly rA-oligo dT) but only at a given protein/template primer ratio. The enzyme is blocked at the amino terminus, thus preventing the automatic sequencing of the protein. The amino acid analysis showed a striking similarity with the animal low molecular weight DNA polymerase beta. The latter observation, as well as the effect of inhibitors (except N-ethylmaleimide which does not inhibit the animal polymerase) indicate that the DNA polymerase described in this work is a plant DNA polymerase very similar to the low molecular weight animal DNA polymerase beta, an enzyme believed to be involved in nuclear DNA repair.
Plant Mol Biol 1990 Sep
PMID:A low molecular weight DNA polymerase from wheat embryos. 210 59


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