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1.5.1.3: dihydrofolate reductase

This is an abbreviated version!
For detailed information about dihydrofolate reductase, go to the full flat file.

Word Map on EC 1.5.1.3

Reaction

5,6,7,8-tetrahydrofolate
+
NADP+
=
7,8-dihydrofolate
 +
NADPH
 +
H+

Synonyms

5,6,7,8-tetrahydrofolate: NADP+ oxidoreductase, 7,8-dihydrofolate reductase, At2g16370, At4g34570, bifunctional dihydrofolate reductase-thymidylate synthase, bifunctional TS-DHFR, BmDHFR, dehydrogenase, tetrahydrofolate, DFR-TS, DFR1, DfrA, DfrB, DHFR, DHFR type IIIC, DHFR-TS, DHFR-TS1, DHFR-TS2, DHFR2, DHFRL1, DHFRLS, dihydrofolate reductase, dihydrofolate reductase-like, dihydrofolate reductase-thymidylate synthase, dihydrofolate reductase:thymidylate synthase, dihydrofolic acid reductase, dihydrofolic reductase, EC 1.5.1.4, ecDHFR, folA, folA3, folic acid reductase, folic reductase, FolM, hDHFR, hDHFR-1, hDHFR-2, HjDHFR, hvDHFR1, hvDHFR2, LAU_0427, LBRM_06_0830, mDHFR, mjDHFR, More, myDHFR, NADPH-dihydrofolate reductase, pcDHFR, PKNH_0509600, ppDHFR, pteridine reductase, pteridine reductase:dihydrofolate reductase, PTR2, R-plasmid-encoded dihydrofolate reductase, R67 DHFR, R67 dihydrofolate reductase, reductase, dihydrofolate, S3DHFR, Smdhfr, Smp 175230, spDHFR, svDHFR, tcptr1, tetrahydrofolate dehydrogenase, THY-1, THY-2, thymidylate synthase-dihydrofolate reductase, thymidylate synthetase-dihydrofolate reductase, Trimethoprim resistance protein, TS-DHFR, WUBG_00817

ECTree

     1 Oxidoreductases
         1.5 Acting on the CH-NH group of donors
             1.5.1 With NAD+ or NADP+ as acceptor
                1.5.1.3 dihydrofolate reductase

General Stability

General Stability on EC 1.5.1.3 - dihydrofolate reductase

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GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
2-mercaptoethanol, 4 mM, stabilizes purified preparation, inclusion in purification steps causes loss of activity
-
4°C, without urea, 50% loss of activity
-
50% residual activity in 4 M urea
Halalkalibacterium halodurans
-
7,8-dihydrofolate and folate protect against ethoxyformic anhydride modification
-
7,8-dihydrofolate protects against heat inactivation
7,8-dihydrofolate protects against inactivation
addition of NaCl (0–500 mM concentration) induces significant structural formation to the enzyme. Protein stability increases depending on NaCl concentration regardless of structural formation, and HjDHFR P1 achieves the same stability as the Escherichia coli enzyme at 750 mM NaCl
AB986558
are more mobile. Betweeen EcDHFR and TmDHFR there is a shift in melting temperature of 26 K
at ionic strengths below the intracellular ion concentration-derived ionic strength in Escherichia coli ( at or below 0.237 M), the DHFR M20 loop tends to adopt open/closed conformations, and rarely an occluded loop state. As the ionic strength exceeds the physiological ionic strength of 0.237 M, the loop tends to adopt a closed/occluded conformation. the solution ionic strength affects the M20 loop stability differently depending on its conformation: High ionic strengths stabilize the occluded conformation more than low ionic strengths, as Ca2+ ions can approach E17 of the M20 loop and stabilize its orientation, whereas they are distal from the E17 at low ionic strengths. Both low and high ionic strengths can stabilize the closed conformation,
bovine serum albumin protects against inactivation
-
bovine serum albumin stabilizes purified enzyme
-
bovine serum albumin, 1 mg/ml, stabilizes during storage of purified enzyme preparation at 5°C
but they do not significantly affect the stability of the open conformation
comparison of the temperature dependence of stability and of flexibility between Thermotoga maritima and Escherichia coli enzymes. The TmDHFR dimer is overall not significantly more rigid than EcDHFR monomer. The TmDHFR protein core and most residues at the dimer interface exhibit smaller fluctuations than in EcDHFR, regions that are opposite to the interface
freezing and thawing, significant loss of activity even in presence of glycerol
-
freezing and thawing, stable to repeated freezing and thawing
-
frozen solutions are not stable for long periods
-
glycerol, 25%, essential for stabilization during purification
-
glycerol, 30%, stabilization of purified preparation
-
isozyme hDHFR-1, loss of more than 80% activity after exposure to 0.2 M KCl for 24 h at 24°C
-
isozyme hDHFR-2 loss of less than 10% activity after exposure to 40 mM KCl for 18 h at 52°C
-
N5-formyltetrahydrofolate, NADP+ and NADPH stabilize and protect against heat inactivation
-
NADPH stabilizes
-
NADPH, protects against ethoxyformic anhydride modification
-
protease inhibitors phenylmethylsulfonyl fluoride and leupeptin are essential for stabilization during purification
-
stabilized by 0.1 mg/ml bovine serum albumin and 1 mM dithiothreitol at 4°C
-
stable in NaCl or KCl up to 0.3 M with slight activation below 0.1 M
Halalkalibacterium halodurans
-
substrates 7,8-dihydrofolate and NADPH stabilize against thermal inactivation
-
the enzyme retains secondary structure at monovalent salt concentrations as low as 0.5 M
unstable to dialysis and lyophilization and concentration by membrane filtration under nitrogen
-
unusually resistant to trypsin
-