Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.1.21 (thymidine kinase)
 7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A human primary lung carcinoma cell line (HPL-R1) established from the tumor biopsy of a lung cancer patient, lacking in cytochrome P1-450 [aryl hydrocarbon (benzo[a]pyrene) hydroxylase (AHH)], was cloned and used to obtain variants deficient in the expression of thymidine-kinase via treatment with 5-bromo-2'-deoxyuridine, and selection for drug resistance phenotype. The variant cell line, precharacterized for thymidine kinase negative phenotype, was transfected with the thymidine kinase gene bearing p R-tk and px1-tk plasmids. Transfections from both the plasmids, demonstrated a frequency of 5.5 X 10(-5). The transfectants showed a 76-100% retention of the transferred phenotype. These data suggest that transfection in variant human cells can approach significant levels of stability observed with rodent cell recipients.
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PMID:Plasmid DNA mediated transfer of the herpes simplex virus thymidine kinase gene to a new bromodeoxyuridine resistant variant of human primary lung carcinoma cells. 283 65

We have demonstrated that the human cytochrome P1-450 gene can be transfected into the AHH-1 human lymphoblastoid cell line using the pHEBo vector and hygromycin selection. The transfected gene was expressed when regulatory sequences derived from the herpes simplex virus thymidine kinase gene were incorporated in appropriate orientations. Gene expression was monitored at the enzyme level using assays for 7-ethoxyresorufin deethylase, 7-ethoxycoumarin deethylase and benzo[a]pyrene hydroxylase activities. Bulk transformed cell populations had 2- to 3-fold more of these enzyme activities compared with control populations. Subclones of the bulk population expressing still higher levels of 7-ethoxyresorufin deethylase activity were also obtained. Expression of the transfected cytochrome P1-450 gene was stable for 20-30 days in the presence of hygromycin B. The transformed cell populations were found to be suitable for use in gene locus mutation assays and the mutagenicity of aflatoxin-B1 and 2-acetylaminofluorene (AAF) were examined. Aflatoxin-B1 was found to be 2-3 times more mutagenic to cells bearing the transfected cytochrome P1-450 activity as compared with control cells. In contrast, no difference in AAF mutagenicity was observed. Analysis of the AAF metabolite profile indicated that cells expressing the transfected cytochrome P1-450 gene produced 8-fold more N- and 7-hydroxy-AAF than control cells. The similarity in mutagenic responses between control cells and cells bearing the transfected cytochrome P1-450 gene may be due to the low deacetylase activity of AHH-1 cells. These observations indicate that this vector and expression system are suitable for introducing novel metabolic activities into the AHH-1 cell line.
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PMID:Transfection of a human cytochrome P-450 gene into the human lymphoblastoid cell line, AHH-1, and use of the recombinant cell line in gene mutation assays. 291 81

Two cDNA clones representing the mRNA coding sequences for mouse cytochromes P1-450 and P3-450 were inserted into the thymidine kinase gene of the wild-type vaccinia virus under the control of the vaccinia virus promoter. Murine and human cells infected with each of the resulting infectious recombinant viruses efficiently expressed their respective P-450 proteins. The newly synthesized protein products are translocated into the microsomes, and their characterization by immunochemical analysis indicates that the sizes of the polypeptides expressed were indistinguishable from their cytochrome P-450 counterparts found in mammalian liver microsomes. Functional analysis of each of the proteins by spectral and enzymatic analysis indicates that the expressed proteins have incorporated heme, and the holoenzymes displayed catalytic activities characteristic of their respective cytochrome P-450 enzymes. Thus, this system can be used to produce properly processed and catalytically active P-450 gene products in a wide variety of cells. The remarkable fidelity of expression and processing of these enzymes suggests that the vaccinia virus recombinants can be used for a wide variety of studies, including analysis of the effects of defined mutations produced in vitro, and directly correlate the structure/activity relationships of the cytochrome P-450 enzymes.
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PMID:Expression of P1-450 and P3-450 DNA coding sequences as enzymatically active cytochromes P-450 in mammalian cells. 303 66