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Literature summary for 2.3.1.30 extracted from

  • Hussain, S.; Ali, V.; Jeelani, G.; Nozaki, T.
    Isoform-dependent feedback regulation of serine O-acetyltransferase isoenzymes involved in L-cysteine biosynthesis of Entamoeba histolytica (2009), Mol. Biochem. Parasitol., 163, 39-47.
    View publication on PubMed

Application

Application Comment Organism
medicine in parasitic protists the L-cysteine biosynthetic pathway is present in contrast to mammals, therefore, enzymes in this pathway are a potential target for chemotherapeutic agents Entamoeba histolytica

Cloned(Commentary)

Cloned (Comment) Organism
PCR-amplification, expression of histidine tagged gene in Escherichia coli BL21 (DE3) pLysS, heat shock transformed Entamoeba histolytica

Inhibitors

Inhibitors Comment Organism Structure
L-cysteine 0.03 mM, 0.5 mM, and 1 mM, strong inhibition, 5-500 microM L-cysteine in the presence of 0.5 mM L-serine inhibits the enzyme remarkably, much less but still significantly in the presence of 3 mM L-serine, 90% inhibition by 0.3 mM L-cysteine with 3 mM L-serine (as found in throphozoite culture), acetyl-CoA concentration influences the inhibitory efficiency as well; 0.5 mM inhibits by 75.3%, no inhibition with 0.03 mM, 1 mM not determined, 20% inhibition by 0.3 mM L-cysteine with 3 mM L-serine (as found in throphozoite culture); 1 mM, weak inhibition, no inhibition by 0.3 mM L-cysteine with 3 mM L-serine (as found in throphozoite culture) or at other L-cysteine concentrations from 10-1000 micoM in the presence of 3 mM L-serine Entamoeba histolytica
additional information no inhibition by 0.03 mM L-cystine, 0.5 mM D-cysteine, DL-homocysteine, DL-homoserine, and oxidized glutathione (reduced glutathione not determined); no inhibition with 0.03 and 0.5 mM L-cystine, 0.5 mM D-cysteine, DL-homocysteine, DL-homoserine, and N-acetyl-L-serine; no inhibition with 0.03 mM L-cystine, 0.5 mM D-cysteine, DL-homocysteine, DL-homoserine, N-acetyl-L-serine, and oxidized glutathione Entamoeba histolytica
N-acetyl-L-cysteine 1 mM weak inhibition; 1 mM, weak inhibition; 1 mM, weak inhibition Entamoeba histolytica
N-acetyl-L-serine 0.5 mM, weak inhibition Entamoeba histolytica
oxidized glutathione 0.5 mM, weak inhibition Entamoeba histolytica
reduced glutathione 0.5 mM weak inhibition; 0.5 mM, weak inhibition Entamoeba histolytica

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.09
-
L-serine SAT2, 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
0.1
-
L-serine SAT2, 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
0.12
-
L-serine 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
0.2
-
acetyl-CoA SAT2, 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica
0.25
-
acetyl-CoA SAT2, 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica
0.55
-
acetyl-CoA 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica

Localization

Localization Comment Organism GeneOntology No. Textmining
cytosol immunofluorescence imaging with antiserum against recombinant protein, and indirectly deduced from missing N-terminal transit peptide Entamoeba histolytica 5829
-
cytosol immunofluorescence imaging with antiserum against recombined protein, and indirectly deduced from missing N-terminal transit peptide Entamoeba histolytica 5829
-

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
34400
-
calculated from sequence, 305 amino acids Entamoeba histolytica
34800
-
calculated from sequence, 311 amino acids Entamoeba histolytica
37000
-
SDS-PAGE, recombinant purified enzyme monomer with 2.6 kDa histidine tag Entamoeba histolytica
37400
-
SDS-PAGE, recombinant purified enzyme monomer with 2.6 kDa histidine tag Entamoeba histolytica
37700
-
calculated from sequence, 336 amino acids Entamoeba histolytica
40000
-
SDS-PAGE, recombinant purified enzyme monomer with 2.6 kDa histidine tag Entamoeba histolytica

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
L-serine + acetyl-CoA Entamoeba histolytica
-
O-acetyl serine + CoA subsequently, cysteine synthase forms L-cysteine from O-acetyl serine ?

Organism

Organism UniProt Comment Textmining
Entamoeba histolytica
-
trophozoites of the clonal strain HM1: IMSS cl 6
-
Entamoeba histolytica Q401L4 trophozoites of the clonal strain HM1: IMSS cl 6
-
Entamoeba histolytica Q401L5 trophozoites of the clonal strain HM1: IMSS cl 6
-

Purification (Commentary)

Purification (Comment) Organism
recombinant cells are centrifuged, pellet washed with PBS, pH 7.4, resuspended in lysis buffer (50 mM Tris-HCl, pH 8.0), incubated, sonicated, centrifuged, Ni2+ -NTA agarose column, washed with buffer (50 mM Tris-HCl, pH 8.0), eluted imidazole, SDS-PAGE, dialyzed with 50 mM Tris-HCl, pH 8.0, storage with 20% glycerol at -80°C Entamoeba histolytica

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
coupling reaction with cysteine synthase, 50 mM Tris-HCl, pH 8.0, 0.4 mM acetyl-CoA, 4 mM L-serine, 5 mM Na2S, 10 mM DTT, recombinant Entamoeba histolytica cysteine synthase 3, 37°C (amount of L-cysteine produced) Entamoeba histolytica
23.42
-
Vmax with acetyl-CoA, 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica
27.77
-
Vmax with L-serine, 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
41.39
-
Vmax with L-serine, 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
65.75
-
Vmax with L-serine, 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
95.1
-
Vmax with acetyl-CoA, 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica
176.8
-
Vmax with acetyl-CoA, 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica

Storage Stability

Storage Stability Organism
-30°C or -80°C, 50 mM Tris-HCl, pH 8.0, 10-20% glycerol in small aliquots, fully active after more than 3 months Entamoeba histolytica

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-serine + acetyl-CoA
-
Entamoeba histolytica O-acetyl serine + CoA subsequently, cysteine synthase forms L-cysteine from O-acetyl serine ?

Subunits

Subunits Comment Organism
monomer
-
Entamoeba histolytica
monomer 1 * 34800, SDS-PAGE Entamoeba histolytica
monomer 1 * 34400, SDS-PAGE Entamoeba histolytica

Synonyms

Synonyms Comment Organism
EhSAT1
-
Entamoeba histolytica
EhSAT2
-
Entamoeba histolytica
EhSAT3
-
Entamoeba histolytica
SAT
-
Entamoeba histolytica
SAT3
-
Entamoeba histolytica
serine acetyltransferase
-
Entamoeba histolytica
serine O-acetyltransferase
-
Entamoeba histolytica

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
14.59
-
acetyl-CoA 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica
17.31
-
L-serine 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
25.52
-
L-serine 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
43.83
-
L-serine 50 mM Tris-HCl, pH 8.0, 0.1 mM acetyl-CoA, 25°C Entamoeba histolytica
58.63
-
acetyl-CoA 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica
117.9
-
acetyl-CoA 50 mM Tris-HCl, pH 8.0, 1 mM L-serine, 25°C Entamoeba histolytica

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.0047
-
L-cysteine competitive inhibition with L-serine but no inhibition with acetyl-CoA, 50 mM Tris-HCl, pH 8.0, 25°C Entamoeba histolytica
0.02779
-
L-cysteine competitive inhibition with L-serine, 50 mM Tris-HCl, pH 8.0, 25°C Entamoeba histolytica
0.0947
-
L-cysteine non-competitive inhibition with acetyl-CoA, 50 mM Tris-HCl, pH 8.0, 0.25 mM acetyl-CoA, 25°C Entamoeba histolytica
0.46
-
L-cysteine mixed inhibition with L-serine but no inhibition with acetyl-CoA, 50 mM Tris-HCl, pH 8.0, 0.5 mM L-serine, 25°C Entamoeba histolytica

pI Value

Organism Comment pI Value Maximum pI Value
Entamoeba histolytica predicted from sequence
-
5.7
Entamoeba histolytica predicted from sequence
-
5.99
Entamoeba histolytica predicted from sequence
-
6.63

IC50 Value

IC50 Value IC50 Value Maximum Comment Organism Inhibitor Structure
additional information
-
0.03 mM inhibits by 97.3%, 0.5 mM by 95.4%, 1 mM by 98.5% Entamoeba histolytica L-cysteine
additional information
-
0.5 mM inhibits by 75.3%, no inhibition with 0.03 mM Entamoeba histolytica L-cysteine
additional information
-
0.5 mM weak inhibition (11.8%) Entamoeba histolytica N-acetyl-L-serine
additional information
-
0.5 mM, 10.3% inhibition Entamoeba histolytica reduced glutathione
additional information
-
0.5 mM, 10.7% inhibition Entamoeba histolytica reduced glutathione
additional information
-
0.5 mM, 19.9% inhibition Entamoeba histolytica oxidized glutathione
additional information
-
1 mM inhibits by 37.8%, no inhibition with 0.5 mM and 0.03 mM Entamoeba histolytica L-cysteine
additional information
-
1 mM weak inhibition (25%) Entamoeba histolytica N-acetyl-L-cysteine
additional information
-
1 mM, 13.3% inhibition Entamoeba histolytica N-acetyl-L-cysteine
additional information
-
1 mM, 27.8% inhibition Entamoeba histolytica N-acetyl-L-cysteine

General Information

General Information Comment Organism
metabolism first step of the L-cysteine biosynthesis pathway Entamoeba histolytica
physiological function the end product L-cysteine is essential for the synthesis of Fe-S proteins and it is necessary for growth, survival, attachment, and anti-oxidation Entamoeba histolytica