Dihydrofolate Reductase human, >=80% (SDS-PAGE), recombinant, expressed in E. coli, >=1 units/mg protein

Synonyms: DHFR; Tetrahydrofolate NADP+ oxidoreductase

MDL No.: MFCD00130955

Purity: >=80% (SDS-PAGE)

Storage: -20C

Enzyme Commission (EC) Number: 1.5.1.3  ( BRENDA | IUBMB )

UNSPSC Code: 12352204

General description: Human DHFR is an 186 amino acid protein with an apparent molecular weight of 25 kDa. It is 30% homologous to the E. coli protein and up to 70% homologous to vertebrate proteins.

Application: Dihydrofolate reductase (DHFR) is a key enzyme in thymidine synthesis. It catalyzes the reduction of dihydrofolate (DHF) to tetrahydrofolate (THF). At a much lower rate, it catayzes the conversion of folate to THF. Since thymidine is a necessary substrate for DNA synthesis, DHFR is a target for anticancer drug development. Methotrexate is the prototype dihydrofolate reductase inhibitor. The enzyme from Sigma has been used in the inhibitory studies of Leishmaniasis donovani pteridine reductase 1 (PTR1). The enzyme has also been used as a positive control to measure the DHFR activity of a protein, MS0308, purified from Mycobacterium smegmatis.

Application: Human dihydrofolate reductase has been used in a study to investigate the stable expression of green fluorescent protein and the targeted disruption of thioredoxin peroxidase-1 gene in Babesia bovis. Human dihydrofolate reductase has also been used in a study to investigate the structural analysis of human dihydrofolate reductase as a binary complex.

Biochem/physiol Actions: The human DHFR gene, as well as DHFR genes in other mammalian species, overcome the inhibitory effects of methotrexate by a mechanism of gene amplification or by amino-acid mutagenesis. Dihydroflate reductase (DHFR) catalyzes the NADPH dependent reduction of dihydrofolate (DHF) to tetrahydrofolate (THF) and, at a much lower rate, the conversion of folate to THF. The reaction product, THF, is an essential cofactor in the conversion of deoxyuridylate (dUMP) to deoxythymidylate (dTMP) by thymidylate synthetase. It is a key enzyme in thymidine synthesis. Therefore, DHFR is a critical enzyme in DNA synthesis and has become a target for drug development and cancer therapy. The variations between DHFR from different sources have enabled the development of species selective DHFR inhibitors, such as trimethoprim (antibacterial and antifungal), pyrimethamine (antiprotozoal), and methotrexate; MTX (antineoplastic, antipsoriatic, and anti-inflammatory).

Biochem/physiol Actions: Km5,6NADPH 0.16 mM 7,8-dihydrofolate 0.03 mM 8-methylpterin 0.13 mMKi7Folate 2.6x10-5 mMMethotrexate 6.1-9x10-9

Physical form: Solution in 10 mM Tris pH 8, 1 mM EDTA, 0.5 mM DTT, 5 muM NADPH, protease inhibitors, and 50% glycerol.

Unit Definition: One unit will convert 1.0 mumole of dihydrofolic acid to tetrahydrofolic acid in 1 minute at pH 7.5 at 22 C.