D6566

Dihydrofolate Reductase human

≥80% (SDS-PAGE), recombinant, expressed in E. coli, ≥1 units/mg protein

• Sinonimo/i: DHFR, Tetrahydrofolate NADP⁺ oxidoreductase
• Classificazione EC (Enzyme Commission): 1.5.1.3 (BRENDA, IUBMB)
• Numero MDL: MFCD00130955
• Codice UNSPSC: 12352204
• NACRES: NA.54

Proprietà

• Ricombinante: expressed in E. coli
• Livello qualitativo: 200
• Saggio: ≥80% (SDS-PAGE)
• Forma fisica: solution
• Attività specifica: ≥1 units/mg protein
• PM: 25 kDa
• Concentrazione: 0.02-0.06 mg/mL
• N° accesso UniProt: P00374
• Condizioni di spedizione: wet ice
• Temperatura di conservazione: −20°C
• Informazioni sul gene: human ... DHFR(1719)

Descrizione

Descrizione generale

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.

Applicazioni

Dihydrofolate Reductase human has been used:

• to investigate the stable expression of green fluorescent protein and the targeted disruption of thioredoxin peroxidase-1 gene in Babesia bovis
• to study the structural analysis of human dihydrofolate reductase as a binary complex
• to study its in vitro kinetic assay for the enzyme inhibition study

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.

Azioni biochim/fisiol

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.

Km^5,6
NADPH 0.16 mM
7,8-dihydrofolate 0.03 mM
8-methylpterin 0.13 mM
Ki⁷
Folate 2.6x10-5 mM
Methotrexate 6.1-9x10^-9

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).

Definizione di unità

One unit will convert 1.0 μmole of dihydrofolic acid to tetrahydrofolic acid in 1 minute at pH 7.5 at 22 °C.

Stato fisico

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

Informazioni sulla sicurezza

• Codice della classe di stoccaggio: 12 - Non Combustible Liquids
• Classe di pericolosità dell'acqua (WGK): WGK 3
• Punto d’infiammabilità (°F): Not applicable
• Punto d’infiammabilità (°C): Not applicable
• Dispositivi di protezione individuale: Eyeshields, Gloves, multi-purpose combination respirator cartridge (US)