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Literature summary for 5.6.2.2 extracted from

  • Sengupta, T.; Mukherjee, M.; Das, A.; Mandal, C.; Das, R.; Mukherjee, T.; Majumder, H.K.
    Characterization of the ATPase activity of topoisomerase II from Leishmania donovani and identification of residues conferring resistance to etoposide (2005), Biochem. J., 390, 419-426.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 5.6.2.2

Cloned(Commentary)

Cloned (Comment) Organism
expression of the His-tagged wild-type and mutant 43 kDa N-terminal domains of the enzyme in Escherichia coli strain BL21(DE3) Leishmania donovani

Protein Variants

Protein Variants Comment Organism
D130G site-directed mutagenesis, reduced ATPase activity compared to the wild-type Leishmania donovani
E155F site-directed mutagenesis, mutation of ATP binding pocket residue in the N-terminal domain, no reduction of ATPase activity compared to the wild-type, altered etoposide binding compared the wild-type Leishmania donovani
G140D site-directed mutagenesis, reduced ATPase activity compared to the wild-type Leishmania donovani
K123A site-directed mutagenesis, mutation of ATP binding pocket residue in the N-terminal domain, no reduction of ATPase activity compared to the wild-type, altered etoposide binding compared the wild-type Leishmania donovani
N65K site-directed mutagenesis, mutation of ATP binding pocket residue in the N-terminal domain, no reduction of ATPase activity compared to the wild-type, no inhibition of the mutant by etoposide Leishmania donovani
N68K site-directed mutagenesis, reduced ATPase activity compared to the wild-type Leishmania donovani
N69K site-directed mutagenesis, mutation of ATP binding pocket residue in the N-terminal domain, reduced ATPase activity compared to the wild-type Leishmania donovani
N96K site-directed mutagenesis, mutation of ATP binding pocket residue in the N-terminal domain, no reduction of ATPase activity compared to the wild-type, no inhibition of the mutant by etoposide Leishmania donovani

Inhibitors

Inhibitors Comment Organism Structure
etoposide traps the enzyme in a covalent complex with DNA, competes with ATP for the N-terminal binding site, binding involves N65, V77, Y127, N128, and N96, inhibits the ATPase activity of the recombinant N-terminal domain by 73% at 0.01 mM, molecular docking analysis, overview Leishmania donovani
luteolin inhibits the ATPase activity of the recombinant N-terminal domain by 57% at 0.02 mM Leishmania donovani
additional information no inhibition of ATPase activity by dihydrobetulinic acid, a pentacyclic triterpenoid derivative Leishmania donovani

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.33
-
 ATP pH 7.5, 30°C, recombinant N-terminal domain, ATPase activity Leishmania donovani

Metals/Ions

Metals/Ions Comment Organism Structure
KCl
-
 Leishmania donovani
Mg2+ required by the N-terminal domain for ATPase activity Leishmania donovani
NaCl
-
 Leishmania donovani

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
43000
-
 x * 43000, recombinant N-terminal enzyme domain, SDS-PAGE Leishmania donovani

Organism

Organism UniProt Comment Textmining
Leishmania donovani Q9XZN0
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant 43 kDa N-terminal domains of the enzyme from Escherichia coli strain BL21(DE3) by nickel affinity and phosphocellulose chromatography Leishmania donovani

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
-
 Leishmania donovani

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
supercoiled DNA + ATP + H2O
-
 Leishmania donovani catenated DNA networks + ADP + phosphate
-
 ?

Subunits

Subunits Comment Organism
? x * 43000, recombinant N-terminal enzyme domain, SDS-PAGE Leishmania donovani
More the recombinant 43 kDa N-terminal domain of the enzyme forms a dimer which is stable in absence or presence of nucleotides, structure modeling, overview Leishmania donovani

Synonyms

Synonyms Comment Organism
Topoisomerase II
-
 Leishmania donovani

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
 ATPase assay at Leishmania donovani

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
 ATPase assay at Leishmania donovani

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.002
-
 etoposide pH 7.5, 30°C, recombinant N-terminal domain, inhibition of ATPase activity Leishmania donovani