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Literature summary for 5.3.1.1 extracted from

  • Gayathri, P.; Banerjee, M.; Vijayalakshmi, A.; Balaram, H.; Balaram, P.; Murthy, M.R.
    Biochemical and structural characterization of residue 96 mutants of Plasmodium falciparum triosephosphate isomerase: active-site loop conformation, hydration and identification of a dimer-interface ligand-binding site (2009), Acta Crystallogr. Sect. D, 65, 847-857.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expression of wild-type and mutant enzymes in TIM-deficient Escherichia coli strain AA200 Plasmodium falciparum

Crystallization (Commentary)

Crystallization (Comment) Organism
wild-type and mutant enzymes in complex with inhibitor 3-phosphoglycerate, 0.003 ml of 10 mg/ml protein in 100 mM triethanolamine-HCl, pH 7.6, are mixed with an equal volume of reservoir solution, the crystallization cocktail contains 0.1 M sodium acetate pH 4.0-5.5 and PEG 1450 varying from 8% to 24% in the reservoir, X-ray diffraction structure determination and analysis at 1.4-2.25 A resolutions Plasmodium falciparum

Protein Variants

Protein Variants Comment Organism
F96H site-directed mutagenesis, the mutant exhibits highly reduced catalytic efficiency and decreased substrate-binding affinity, as well as reduced sensitivity to inhibitor 3-phosphoglycerate, compared to the wild-type enzyme Plasmodium falciparum
F96S site-directed mutagenesis, the mutant exhibits highly reduced catalytic efficiency and decreased substrate-binding affinity, as well as reduced sensitivity to inhibitor 3-phosphoglycerate, compared to the wild-type enzyme Plasmodium falciparum
F96W site-directed mutagenesis, the mutant exhibits reduced catalytic efficiency and decreased substrate-binding affinity, as well as reduced sensitivity to inhibitor 3-phosphoglycerate, compared to the wild-type enzyme. The wild-type enzyme shows a loop-open state for 3-phosphoglycerate binding at the active site, while the mutant F96W shows both open and closed states Plasmodium falciparum
additional information decrease in ligand affinity in F96 mutants can be a consequence of differences in the water network connecting residue 96 to Ser73 in the vicinity of the active site Plasmodium falciparum

Inhibitors

Inhibitors Comment Organism Structure
3-phosphoglycerate binding at the active site with the dimer-interface site showing strong electrostatic anchoring of the phosphate group involving the Arg98 and Lys112 residues of TIM, comparisons of binding structures at the interface, overview Plasmodium falciparum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.39
-
D-glyceraldehyde 3-phosphate pH 7.6, wild-type enzyme Plasmodium falciparum
1.2
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96W Plasmodium falciparum
2.18
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96S Plasmodium falciparum
2.62
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96H Plasmodium falciparum

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
D-Glyceraldehyde 3-phosphate Plasmodium falciparum
-
Glycerone phosphate
-
?

Organism

Organism UniProt Comment Textmining
Plasmodium falciparum Q07412
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes from TIM-deficient Escherichia coli strain AA200 by gel filtration and ion-exchange chromatography Plasmodium falciparum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-Glyceraldehyde 3-phosphate
-
Plasmodium falciparum Glycerone phosphate
-
?
additional information Plasmodium falciparum TIM is unique in possessing a Phe residue at position 96 in place of the conserved Ser that is found in TIMs from the majority of other organisms Plasmodium falciparum ?
-
?

Synonyms

Synonyms Comment Organism
TIM
-
Plasmodium falciparum
Triosephosphate isomerase
-
Plasmodium falciparum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
185
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96H Plasmodium falciparum
620
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96S Plasmodium falciparum
2466
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96W Plasmodium falciparum
42330
-
D-glyceraldehyde 3-phosphate pH 7.6, wild-type enzyme Plasmodium falciparum

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.6
-
assay at Plasmodium falciparum

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
1.9
-
3-phosphoglycerate pH 7.6, wild-type enzyme Plasmodium falciparum
5
-
3-phosphoglycerate above, pH 7.6, mutants F96W, F96S, and F96H Plasmodium falciparum

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
28.5
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96S Plasmodium falciparum
70.5
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96H Plasmodium falciparum
2050
-
D-glyceraldehyde 3-phosphate pH 7.6, mutant F96W Plasmodium falciparum
10850
-
D-glyceraldehyde 3-phosphate pH 7.6, wild-type enzyme Plasmodium falciparum