Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.5.1.3 (dihydrofolate reductase)
 5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

DNA fragments of the R factor R388 which renders E. coli resistant to trimethoprim by inducing a trimethoprim resistant dihydrofolate reductase (Amyes and Smith, 1974) were inserted into plasmids and screened for the expression of the trimethoprim resistance gene. By means of a two step deletion procedure a 1770 bp EcoRI/BamH1 fragment was isolated which conferred drug resistance and which was found to induce the synthesis of the same dihydrofolate reductase as the parental R factor. Gene dosage experiments indicated that the induction was due to the presence of a dihydrofolate reductase structural gene on the 1770 bp fragment. The gene could be assigned to a segment which was less than 1200 bp long. The 1770 bp fragment and a recombinant plasmid consisting of pSF2124 and part of R388 were mapped with several restriction nucleases. The R factor induced enzyme was partially purified from a strain carrying a multicopy recombinant plasmid into which the 1770 bp fragment was inserted and which induced high levels of dihydrofolate reductase. The enzyme was found to be stable at 100 degrees. Some aspects of the synthesis of dihydrofolate reductase are discussed.
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PMID:Isolation of a small DNA fragment carrying the gene for a dihydrofolate reductase from a trimethoprim resistance factor. 36 38

The construction and analysis of bacterial plasmids that contain and phenotypically express a mammalian genetic sequence are described. Such plasmids specify a protein that has enzymatic properties, immunological reactivity and molecular size characteristic of the mouse dihydrofolate reductase, and render host cells resistant to the antimetabolic drug trimethoprim.
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PMID:Phenotypic expression in E. coli of a DNA sequence coding for mouse dihydrofolate reductase. 36 74

Dihydrofolate reductase, specified by the type II plasmid of a trimethoprim-resistant Escherichia coli, was purified 40-fold to homogeneity using a combination of gel filtration, DEAE-Sephacel chromatography, and hydrophobic chromatography. The final product shows a single protein band on polyacrylamide gel electrophoresis and has a specific activity of 1.0 unit/mg. The molecular weight of the purified enzyme is 36,000 as determined both by gel filtration and Ferguson analysis of polyacrylamide gel electrophoresis. In contrast, a single polypeptide with a molecular weight of 8,500 was observed on sodium dodecyl sulfate-gel electrophoresis. These experiments suggest that, unlike any bacteria or vertebrate dihydrofolate reductase previously examined, the type II R plasmid reductase is a tetramer composed of four identical subunits. A partial amino acid sequence determination shows no heterogeneity of the subunits and also no clear homology with any reductase sequence previously reported.
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PMID:R plasmid dihydrofolate reductase with subunit structure. 37 28

The amino acid sequence of a trimethoprim-resistant dihydrofolate reductase (EC 1.5.1.3) specified by the R-plasmid R67 is described. The sequence was deduced from automatic and manual sequence analysis of the intact protein, the fragments produced by cyanogen bromide cleavage, and peptides derived from the largest cyanogen bromide fragment by digestion with trypsin, Staphylococcus aureus V8 proteus, chymotrypsin, and Lysobacter enzymogenes alpha-lytic protease. The complete sequence comprises 78 residues in a single polypeptide chain of molecular weight 8444. No evidence of heterogeneity was obtained, indicating that all subunits of the native enzyme are identical. Comparison of the sequence with that of all known dihydrofolate reductases shows no significant sequence homology.
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PMID:The amino acid sequence of the trimethoprim-resistant dihydrofolate reductase specified in Escherichia coli by R-plasmid R67. 38 58

The paper presents a survey of literature concerned with the possibility of expression of plasmid-clones genes from eukaryotic organisms in bacteria cells. Studies on bacterial synthesis of somatostatin, human insulin, hormone of rat growth and proteins: chicken ovalbumin and mouse dihydrofolate reductase are discussed.
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PMID:[Expression of eukaryotic genes in Escherichia coli cells]. 38 36

Two clones from the Clarke-Carbon Escherichia coli colony bank were resistant to inhibition by trimethoprim, a potent inhibitor of dihydrofolate reductase. Both clones had elevated levels of dihydrofolate reductase. Furthermore, trimethoprim resistance and elevated enzyme levels were associated with ColE1 plasmids that carried DNA from the trkC ksgA pdxA region of the E. coli chromosome. Plasmid pLC1437a was shown by two criteria to carry the structural gene for dihydrofolate reductase: 1) A partial diploid containing plasmid pLC1437a produced a kinetically-recognizable dihydrofolate reductase that was not present in the parent haploid strain. 2) Plasmid pLC1437a coded for dihydrofolate reductase in vitro. A 1,000 base pair fragment of plasmid pLC1437a containing fol was used as a probe to measure fol mRNA in a mutant strain isolated by Sheldon and Brenner (Molec. gen. Genet. 147, 91-97, 1976). The mutation in this strain, which results in constitutively-high levels of dihydrofolate reductase and in the inability of the strain to grow at 42 degrees C, is cis dominant (Sheldon and Brenner, 1976). The results of kinetic hybridization and pulse-labeling experiments indicated that the regulatory mutant produced elevated levels of dihydrofolate reductase in response to an increased rate of synthesis of fol mRNA.
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PMID:Regulation of dihydrofolate reductase synthesis in Escherichia coli. 39 Mar 4

Four hr after either a single injection or continuous infusion of methotrexate (MTX) plus purified [3',5',9(n)-3H]MTX in cynomolgus or rhesus monkeys, 80 to 98% of the 3H radioactivity present in the plasma was found not to represent intact MTX. The percentage of 3H-containing MTX products in the urine after 4 hr was considerably less, although more variable. This variability seemed to be related to variability in the amount of the total dose excreted. Non-MTX products were also found in selected tissues and the percentage of intact MTX found 4 hr after i.v. injection varied from 2 to 26%. The percentage of intact MTX was routinely measured by comparing the values obtained using the dihydrofolate reductase assay with values based on the specific activity of [3',5',9(n)-3H]MTX. Results obtained by diethylaminoethyl column chromatography on a few samples, however, showed good agreement with results from the reductase assay. [3',5',9(n)-3H]MTX products appeared in peaks eluting from the diethylaminoethyl column both earlier and later than the MTX peak, with the earlier peaks being present in only small amounts in the urine. After continuous i.v. infusion, only 2% or less of the radioactivity found in the cerebrospinal fluid after 4 hr represented intact MTX, with the remaining radioactivity eluting much earlier than MTX. In contrast, after direct injection into the left lateral ventricel, all the 3H radioactivity in both cerebrospinal fluid and brain tissue represented intact MTX for up to 4 hr after injection. The appearance of MTX products in the plasma and selected tissues of these primates a short time after i.v. injection is compared to other work in experimental animals and man and suggests a greater metabolism of MTX than was previously suspected.
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PMID:Distribution and degradation of [3H]methotrexate after intravenous and cerebral intraventricular injection in primates. 40 70

Humans and rhesus monkeys receiving high-dose methotrexate (MTX) (greater than 50 mg/kg) excrete significant quantities of the metabolite, 7-hydroxy-MTX. This metabolite, though 200-fold less potent than MTX as an inhibitor of mammalian dihydrofolate reductase, is of very limited aqueous solubility, and thus may contribute to the renal toxicity of the high-dose regimen. The metabolite was not observed in previous pharmacologic studies in which conventional doses of MTX were administered (less than 10 mg/kg).
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PMID:Dose-dependent metabolism of methotrexate in man and rhesus monkeys. 40 97

A series of 8-alkyl-7,8,-dihydromethotrexate analogues was prepared by direct alkylation of 7,8-dihydromethotrexate, after pilot studies were performed with simpler pteridines. These compounds are tested for in vitro inhibitory activity against Lactobacillus casei and as enzyme inhibitors against dihydrofolate reductase and thymidylate synthetase derived from this organism. All of the analogues were less inhibitory toward dihydrofolate reductase than was methotrexate but were more inhibitory toward thymidylate synthetase. The analogues were also evaluated for in vitro inhibitory activity against the CCRF-CEM human lymphoblastic leukemia cells. In vivo against the L-1210 leukemia in mice, several of the analogues exhibited some antileukemic activity.
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PMID:Methotrexate analogues. 9. Synthesis and biological properties of some 8-alkyl-7,8-dihydro analogues. 40 42

The diastereoisomers of 5,10-methylene 5,6,7,8-tetrahydropteroyl-D-glutamate were resolved and tested as substrates and inhibitors of Lactobacillus casei thymidylate synthetase. No activity was observed. The compounds were neither growth factors nor inhibitors for Lactobacillus casei, Streptococcus faecium, or Pediococcus cerevisiae. 7,8-Dihydropteroyl-D-glutamate is 50% as active as 7,8-dihydropteroyl-L-glutamate (dihydrofolate) as a substrate for L. casei dihydrofolate reductase.
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PMID:Diastereoisomers of 5,10-methylene-5,6,7,8-tetrahydropteroyl-D-glutamic acid. 41 Sep 32


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