[185] A new preparation of dihydrofolic acid

Zakrzewski, Sigmund F.; Sansone, Annette M.

doi:10.1016/s0076-6879(71)18142-6

Cited by 7 publications

(3 citation statements)

References 2 publications

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“…Other folates used as standards for TEAE-cellulose chromatography and high-pressure liquid chromatography (HPLC) were prepared as follows: dl-5-CH3-H4PteGlu was synthesized by the method of Gupta & Huennekens (1967) and purified according to Horne et al (1978). H2PteGlu was synthesized byZn plus NaOH reduction of PteGlu (Zakrewski & Sansone, 1971). H4PteGlu was made by catalytic hydrogenation (Blakely, 1957).…”

Section: Methodsmentioning

confidence: 99%

Purification and partial characterization of rat liver folate binding protein: cytosol I

Cook¹,

Wagner²

1982

Biochemistry

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The high molecular weight folate binding protein of rat liver cytosol has been purified to apparent homogeneity. Purification was achieved by using a combination of gel filtration, O-(diethylaminoethyl)cellulose chromatography, and affinity chromatography. This folate binding protein was initially identified during purification by an in vivo labeling procedure involving intraperitoneal injection of [3H]folic acid prior to sacrifice and subsequently by its ability to bind naturally reduced [3H]folate polyglutamates in vitro. A molecular weight of 210 000 was estimated by gel chromatography. This is distinct from the trifunctional formyl-methenyl-methylene synthetase of rat liver which has a molecular weight of 225 000. Sodium dodecyl sulfate electrophoresis revealed a single band with a molecular weight of about 100 000 which suggests the native protein is composed of two identical subunits. The partially purified protein contains bound tetrahydropteroylpentaglutamate.

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Section: Methodsmentioning

confidence: 99%

Purification and partial characterization of rat liver folate binding protein: cytosol I

Cook¹,

Wagner²

1982

Biochemistry

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“…Dihydrofolate (DHF) was produced from folate by the method of Zakrzewski and Sansone 9 ) with some modifications. One hundred and fifty grams of folate was dissolved in I liter of 3 M NaOH and the pH was adjusted with HCI to 13.5.…”

Section: Methodsmentioning

confidence: 99%

Production of Dihydrofolate Reductase by ClonedEscherichia coliand Its Application to Asymmetric Synthesis ofl-Leucovorin

Ohshiro

Kuge²,

Igarashi³

et al. 1992

Bioscience, Biotechnology, and Biochemistry

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“…10-CH3-folate, as obtained above or from the alkaline hydrolysis of methotrexate, was purified by DEAE-cellulose chromatography and reduced catalytically to yield 10-CH3-H4folate (Gupta & Huennekens, 1967). H2folate (Sigma) was purified by DEAE-cellulose column chromatography (Zakrewski & Sansone, 1971). The racemic mixture (±) of H4folate was obtained by catalytic hydrogenation of folic acid in glacial acetic acid as described by Hatefi et al (1960).…”

Section: Methodsmentioning

confidence: 99%

Fluorine-19 nuclear magnetic resonance characterization of ternary complexes of folate derivatives, 5-fluorodeoxyuridylate and Lactobacillus casei thymidylate synthetase

Ca¹,

Pd²,

Rb³

1981

Biochemistry

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Numerous biochemical techniques have been employed to characterize the covalent inhibitory ternary complex of thymidylate synthetase consisting of enzyme, 5-fluorodeoxyuridylate, and 5,10-methylenetetrahydrofolate. 19F NMR studies of this covalent ternary complex reveal a single, broad resonance centered at 12.7 ppm to higher shielding of free nucleotide, while the 5-fluorodexyuridylate-enzyme binary complex exhibits two resonances to higher shielding of free nucleotide, one at 1.4 ppm representing noncovalently bound ligand and the other at 34.5 ppm indicative of covalently bound 5,6-dihydro-5-fluorodeoxyuridylate. In order to follow the transformation of the latter binary complex to a ternary complex, we have employed 19F NMR to profile changes in the environment of the nucleotide which result from the interaction of folates with the coenzyme binding site. At low molar excesses of folates (5-fold), the effects observed in the 19F NMR spectrum fall into three major classes. (1) 5-Methyltetrahydrofolate exhibited a weak interaction with the binary complex. (2) Methotrexate and aminopterin, antifolate drugs, were observed to increase the exchange rate among the species detected in the 19F NMR spectrum of the binary complex. (3) Folate, dihydrofolate, and a series of tetrahydrofolate derivatives were found to shift the equilibrium of the binary complex toward the covalent 5,6-dihydro-5-fluorodeoxyuridylate-enzyme complex. With the latter folates the chemical shifts for the covalent species of these ternary complexes were found in the range of 35-40 ppm to higher shielding and are interpreted to reflect subtle differences in the strength and steric nature of the interaction of the folate ligand with the binary complex. These data illustrate that the latter folates promote the conversion of the enzyme-bound nucleotide to a species which would be poised to form the second covalent bond of the ternary complex, namely the linkage of the methylene group of the coenzyme with carbon 5 of the nucleotide.

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[185] A new preparation of dihydrofolic acid

Cited by 7 publications

References 2 publications

Purification and partial characterization of rat liver folate binding protein: cytosol I

Purification and partial characterization of rat liver folate binding protein: cytosol I

Production of Dihydrofolate Reductase by ClonedEscherichia coliand Its Application to Asymmetric Synthesis ofl-Leucovorin

Fluorine-19 nuclear magnetic resonance characterization of ternary complexes of folate derivatives, 5-fluorodeoxyuridylate and Lactobacillus casei thymidylate synthetase

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