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Literature summary for 6.2.1.13 extracted from

  • Pineda, E.; Vazquez, C.; Encalada, R.; Nozaki, T.; Sato, E.; Hanadate, Y.; Nequiz, M.; Olivos-Garcia, A.; Moreno-Sanchez, R.; Saavedra, E.
    Roles of acetyl-CoA synthetase (ADP-forming) and acetate kinase (PPi-forming) in ATP and PPi supply in Entamoeba histolytica (2016), Biochim. Biophys. Acta, 1860, 1163-1172 .
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
recombinant overexpression of His-tagged enzyme in Escherichia coli strain BL21(DE3), quantitative RT-PCR enzyme expression analysis Entamoeba histolytica

Protein Variants

Protein Variants Comment Organism
additional information gradual ACS gene silencing (49-93%) significantly decreases the acetate flux without affecting the levels of glycolytic metabolites and ATP in trophozoites. Amoebae lacking ACS activity are unable to reestablish the acetyl-CoA/CoA ratio after an oxidative stress challenge Entamoeba histolytica

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics, the amoebal ACS displays hyperbolic kinetics for its substrates acetyl-CoA, Pi and ADP and its products acetate, ATP and CoA Entamoeba histolytica
0.13
-
acetyl-CoA at pH 7.0 and 37°C Entamoeba histolytica
0.14
-
acetyl-CoA pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
0.14
-
acetyl-CoA at pH 6.0 and 37°C Entamoeba histolytica
0.2
-
CoA pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
0.2
-
CoA at pH 7.0 and 37°C Entamoeba histolytica
0.3
-
ATP pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
0.3
-
ATP at pH 7.0 and 37°C Entamoeba histolytica
0.4
-
CoA at pH 6.0 and 37°C Entamoeba histolytica
0.6
-
ATP at pH 6.0 and 37°C Entamoeba histolytica
0.8
-
acetate pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
0.8
-
acetate at pH 7.0 and 37°C Entamoeba histolytica
0.9
-
ADP at pH 7.0 and 37°C Entamoeba histolytica
1.3
-
acetate at pH 6.0 and 37°C Entamoeba histolytica
1.8
-
phosphate at pH 7.0 and 37°C Entamoeba histolytica
1.9
-
ADP pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
1.9
-
ADP at pH 6.0 and 37°C Entamoeba histolytica
2.7
-
phosphate pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
2.7
-
phosphate at pH 6.0 and 37°C Entamoeba histolytica

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Entamoeba histolytica

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ADP + phosphate + acetyl-CoA Entamoeba histolytica
-
ATP + acetate + CoA
-
r
ATP + acetate + CoA Entamoeba histolytica
-
ADP + phosphate + acetyl-CoA
-
r

Organism

Organism UniProt Comment Textmining
Entamoeba histolytica Q9NAT4
-
-

Purification (Commentary)

Purification (Comment) Organism
Co2+ affinity column chromatography Entamoeba histolytica
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by cobalt affinity chromatography Entamoeba histolytica

Source Tissue

Source Tissue Comment Organism Textmining
trophozoite
-
Entamoeba histolytica
-

Storage Stability

Storage Stability Organism
-20°C, 50% (v/v) glycerol, 4 weeks, no loss of activity Entamoeba histolytica

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ADP + phosphate + acetyl-CoA
-
Entamoeba histolytica ATP + acetate + CoA
-
r
ATP + acetate + CoA
-
Entamoeba histolytica ADP + phosphate + acetyl-CoA
-
r
additional information metabolites of both reaction directions are determined by gas chromatography Entamoeba histolytica ?
-
?

Subunits

Subunits Comment Organism
homodimer 2 * 79000, SDS-PAGE Entamoeba histolytica

Synonyms

Synonyms Comment Organism
Acetyl-CoA synthetase (ADP-forming)
-
Entamoeba histolytica
ACS
-
Entamoeba histolytica
ADP-forming acetyl-CoA synthetase
-
Entamoeba histolytica

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Entamoeba histolytica

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
34
-
ADP pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
34
-
phosphate pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
34
-
acetyl-CoA pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
34
-
acetyl-CoA at pH 6.0 and 37°C Entamoeba histolytica
35
-
acetate at pH 6.0 and 37°C Entamoeba histolytica
46
-
ATP pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
46
-
CoA pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
46
-
acetate pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
46
-
acetate at pH 7.0 and 37°C Entamoeba histolytica
55
-
acetyl-CoA at pH 7.0 and 37°C Entamoeba histolytica

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6
-
forward reaction Entamoeba histolytica
7
-
reverse reaction Entamoeba histolytica
8
-
-
Entamoeba histolytica

pH Range

pH Minimum pH Maximum Comment Organism
6 7 at pH 6.0-7.0, the recombinant enzyme shows 15–60% of the maximum activity Entamoeba histolytica

Cofactor

Cofactor Comment Organism Structure
ATP
-
Entamoeba histolytica

General Information

General Information Comment Organism
malfunction amoebae lacking enzyme activity are unable to reestablish the acetyl-CoA/CoA ratio after an oxidative stress challenge Entamoeba histolytica
malfunction gradual ACS gene silencing (49-93%) significantly decreases the acetate flux without affecting the levels of glycolytic metabolites and ATP in trophozoites. Amoebae lacking ACS activity are unable to reestablish the acetyl-CoA/CoA ratio after an oxidative stress challenge Entamoeba histolytica
metabolism the enzyme is related to the phosphate-dependent fermentative glycolysis in Entamoeba histolytica. The end-products of carbohydrate catabolism in the parasite are ethanol and acetate, produced by the bifunctional aldehyde-alcohol dehydrogenase (ADHE) and ACS, respectively, overview Entamoeba histolytica
metabolism the main role of the enzyme in the parasite energy metabolism is to recycle CoA for glycolysis to proceed under aerobic conditions Entamoeba histolytica

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
12
-
phosphate at pH 6.0 and 37°C Entamoeba histolytica
12
-
phosphate pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
17
-
ADP at pH 6.0 and 37°C Entamoeba histolytica
17
-
ADP pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
26
-
acetate at pH 6.0 and 37°C Entamoeba histolytica
32
-
phosphate at pH 7.0 and 37°C Entamoeba histolytica
58
-
acetate at pH 7.0 and 37°C Entamoeba histolytica
58
-
acetate pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
59
-
ATP at pH 6.0 and 37°C Entamoeba histolytica
76
-
ADP at pH 7.0 and 37°C Entamoeba histolytica
90
-
CoA at pH 6.0 and 37°C Entamoeba histolytica
144
-
ATP at pH 7.0 and 37°C Entamoeba histolytica
144
-
ATP pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
213
-
CoA at pH 7.0 and 37°C Entamoeba histolytica
213
-
CoA pH 6.0, 37°C, recombinant enzyme Entamoeba histolytica
239
-
acetyl-CoA at pH 6.0 and 37°C Entamoeba histolytica
239
-
acetyl-CoA pH 7.0, 37°C, recombinant enzyme Entamoeba histolytica
458
-
acetyl-CoA at pH 7.0 and 37°C Entamoeba histolytica