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Literature summary extracted from

  • Eadsforth, T.C.; Cameron, S.; Hunter, W.N.
    The crystal structure of Leishmania major N5,N10-methylenetetrahydrofolate dehydrogenase/cyclohydrolase and assessment of a potential drug target (2012), Mol. Biochem. Parasitol., 181, 178-185.
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
3.5.4.9 drug development the bifunctional enzyme is a target for design of parasite-specific inhibitors as structure-based drugs Leishmania major

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.5.1.5 expressed in Escherichia coli BL21(DE3) cells Leishmania major
3.5.4.9 expression of the His-tagged enzyme in Escherichia coli Leishmania major

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.5.1.5 hanging drop vapor diffusion method, using 20% (w/v) PEG 3350 and 0.2 M ammonium acetate Leishmania major
3.5.4.9 purified recombinant apo-DHCH, hanging drop vapor diffusion, mixing of 0.002 ml 9 mg/ml protein in 50 mM Tris–HCl, 100 mM NaCl, pH 7.5, with 0.002 ml reservoir solution containing 20% PEG 3350 and 0.2 M ammonium acetate, equilibration against 0.07 ml of reservoir solution at 22°C, 3 days, X-ray diffraction structure determination and analysis at 2.7 A resolution, modelling Leishmania major

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.5.1.5 LY354899 i.e. 5,6,7,8-tetrahydro N5,N10-carbonylfolic acid Leishmania major
3.5.4.9 5,6,7,8-tetrahydro-N5,N10-carbonylfolic acid i.e. LY354899 Leishmania major
3.5.4.9 di-2'-deoxyadenosine 5',5''-P1,P5,alpha,beta-methylene-delta,epsilon-methylene-pentaphosphate-gamma-borano
-
Leishmania major
3.5.4.9 di-2'-deoxyadenosine 5',5''-P1,P5,alpha,beta-methylene-gamma,delta-methylene-tetraphosphate
-
Leishmania major
3.5.4.9 diadenosine 5',5''-P1,P5,alpha,beta-methylene-delta,epsilon-methylene-pentaphosphate-gamma-borano
-
Leishmania major
3.5.4.9 P1,P4-bis(5'-adenosyl)-alpha,beta-gamma,delta-bismethylene-tetraphosphate
-
Leishmania major

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.5.1.5 31900
-
2 * 31900, calculated from amino acid sequence Leishmania major
1.5.1.5 65000
-
gel filtration Leishmania major
3.5.4.9 65000
-
recombinant detagged enzyme, gel filtration Leishmania major

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.5.1.5 5,10-methylenetetrahydrofolate + NADP+ Leishmania major
-
5,10-methenyltetrahydrofolate + NADPH + H+
-
r
1.5.1.5 5,10-methylenetetrahydrofolate + NADP+ Leishmania major Fridlin
-
5,10-methenyltetrahydrofolate + NADPH + H+
-
r
3.5.4.9 additional information Leishmania major a bifuncitonal enzyme exhibiting N5,N10-methylenetetrahydrofolate dehydrogenase, EC 1.5.1.5, and N5,N10-methenyltetrahydrofolate cyclohydrolase activities ?
-
?
3.5.4.9 additional information Leishmania major Fridlin a bifuncitonal enzyme exhibiting N5,N10-methylenetetrahydrofolate dehydrogenase, EC 1.5.1.5, and N5,N10-methenyltetrahydrofolate cyclohydrolase activities ?
-
?
3.5.4.9 N5,N10-methenyltetrahydrofolate + H2O Leishmania major
-
N10-formyltetrahydrofolate
-
?
3.5.4.9 N5,N10-methenyltetrahydrofolate + H2O Leishmania major Fridlin
-
N10-formyltetrahydrofolate
-
?

Organism

EC Number Organism UniProt Comment Textmining
1.5.1.5 Leishmania major Q4Q9F9
-
-
1.5.1.5 Leishmania major Fridlin Q4Q9F9
-
-
3.5.4.9 Leishmania major Q4Q9F9
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.5.1.5 HisTrap column chromatography and Superdex 200 gel filtration Leishmania major
3.5.4.9 recombinant His-tagged enzyme by nickel affinity chromatography, cleavage of the His-tag by TEV protease, His affinity chromatography, and gel filtration, to over 95% purity Leishmania major

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.5.1.5 5,10-methylenetetrahydrofolate + NADP+
-
Leishmania major 5,10-methenyltetrahydrofolate + NADPH + H+
-
r
1.5.1.5 5,10-methylenetetrahydrofolate + NADP+
-
Leishmania major Fridlin 5,10-methenyltetrahydrofolate + NADPH + H+
-
r
3.5.4.9 additional information a bifuncitonal enzyme exhibiting N5,N10-methylenetetrahydrofolate dehydrogenase, EC 1.5.1.5, and N5,N10-methenyltetrahydrofolate cyclohydrolase activities Leishmania major ?
-
?
3.5.4.9 additional information a bifuncitonal enzyme exhibiting N5,N10-methylenetetrahydrofolate dehydrogenase, EC 1.5.1.5, and N5,N10-methenyltetrahydrofolate cyclohydrolase activities Leishmania major Fridlin ?
-
?
3.5.4.9 N5,N10-methenyltetrahydrofolate + H2O
-
Leishmania major N10-formyltetrahydrofolate
-
?
3.5.4.9 N5,N10-methenyltetrahydrofolate + H2O
-
Leishmania major Fridlin N10-formyltetrahydrofolate
-
?

Subunits

EC Number Subunits Comment Organism
1.5.1.5 homodimer 2 * 31900, calculated from amino acid sequence Leishmania major
3.5.4.9 homodimer
-
Leishmania major

Synonyms

EC Number Synonyms Comment Organism
1.5.1.5 DHCH
-
Leishmania major
1.5.1.5 N5,N10-methylenetetrahydrofolate dehydrogenase/cyclohydrolase bifunctional enzyme Leishmania major
3.5.4.9 DHCH
-
Leishmania major
3.5.4.9 N5,N10-methylenetetrahydrofolate dehydrogenase/cyclohydrolase
-
Leishmania major
3.5.4.9 N5,N10-methylenetetrahydrofolate dehydrogenase/N5,N10-methenyltetrahydrofolate cyclohydrolase
-
Leishmania major

Cofactor

EC Number Cofactor Comment Organism Structure
1.5.1.5 NADP+
-
Leishmania major

Ki Value [mM]

EC Number Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
1.5.1.5 0.000105
-
LY354899 pH and temperature not specified in the publication Leishmania major
3.5.4.9 0.009
-
di-2'-deoxyadenosine 5',5''-P1,P5,alpha,beta-methylene-delta,epsilon-methylene-pentaphosphate-gamma-borano with NPP1, pH 7.3, 37°C Leishmania major
3.5.4.9 0.013
-
P1,P4-bis(5'-adenosyl)-alpha,beta-gamma,delta-bismethylene-tetraphosphate with NPP1, pH 7.3, 37°C Leishmania major
3.5.4.9 0.018
-
5,6,7,8-tetrahydro-N5,N10-carbonylfolic acid , pH 7.3, 37°C Leishmania major
3.5.4.9 0.02
-
diadenosine 5',5''-P1,P5,alpha,beta-methylene-delta,epsilon-methylene-pentaphosphate-gamma-borano with NPP1, pH 7.3, 37°C Leishmania major
3.5.4.9 0.051
-
di-2'-deoxyadenosine 5',5''-P1,P5,alpha,beta-methylene-gamma,delta-methylene-tetraphosphate with NPP1, pH 7.3, 37°C Leishmania major

IC50 Value

EC Number IC50 Value IC50 Value Maximum Comment Organism Inhibitor Structure
3.5.4.9 0.013
-
with NPP1, pH 7.3, 37°C Leishmania major di-2'-deoxyadenosine 5',5''-P1,P5,alpha,beta-methylene-delta,epsilon-methylene-pentaphosphate-gamma-borano
3.5.4.9 0.033
-
with NPP1, pH 7.3, 37°C Leishmania major P1,P4-bis(5'-adenosyl)-alpha,beta-gamma,delta-bismethylene-tetraphosphate
3.5.4.9 0.05
-
with NPP1, pH 7.3, 37°C Leishmania major di-2'-deoxyadenosine 5',5''-P1,P5,alpha,beta-methylene-gamma,delta-methylene-tetraphosphate
3.5.4.9 0.063
-
with NPP1, pH 7.3, 37°C Leishmania major diadenosine 5',5''-P1,P5,alpha,beta-methylene-delta,epsilon-methylene-pentaphosphate-gamma-borano

General Information

EC Number General Information Comment Organism
3.5.4.9 metabolism three enzyme activities in the protozoan Leishmania major, namely N5,N10-methylenetetrahydrofolate dehydrogenase/N5,N10-methenyltetrahydrofolate cyclohydrolase and N10-formyltetrahydrofolate ligase produce the essential intermediate N10-formyltetrahydrofolate, folate metabolism pathway, overview Leishmania major
3.5.4.9 additional information active site structure, overview Leishmania major