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Literature summary for 2.3.1.5 extracted from

  • Sikora, A.L.; Frankel, B.A.; Blanchard, J.S.
    Kinetic and chemical mechanism of arylamine N-acetyltransferase from Mycobacterium tuberculosis (2008), Biochemistry, 47, 10781-10789.
    View publication on PubMedView publication on EuropePMC

Application

Application Comment Organism
medicine TBNAT remains a potential drug target for the treatment of tuberculosis Mycobacterium tuberculosis

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli as a His-tagged fusion protein Mycobacterium tuberculosis
into the pET28a+ vector for expression in Escherichia coli BL21DE3 cells Mycobacterium tuberculosis

Inhibitors

Inhibitors Comment Organism Structure
iodoacetamide incubation of TBNAT with iodoacetamide results in a time-dependentloss of enzymatic activity Mycobacterium tuberculosis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.32
-
3-Amino-4-hydroxybenzoic acid
-
Mycobacterium tuberculosis
0.61
-
hydralazine
-
Mycobacterium tuberculosis
0.9
-
4-amino-3-hydroxybenzoic acid
-
Mycobacterium tuberculosis
1.9
-
2-amino-4-chlorophenol
-
Mycobacterium tuberculosis
2 3 aniline
-
Mycobacterium tuberculosis
3.06
-
2-amino-4-methylphenol
-
Mycobacterium tuberculosis
5
-
p-Aminobenzoic acid
-
Mycobacterium tuberculosis
5.79
-
2-Aminophenol
-
Mycobacterium tuberculosis
6
-
4-chloroaniline
-
Mycobacterium tuberculosis
11
-
4-methylaniline
-
Mycobacterium tuberculosis
14
-
anisidine
-
Mycobacterium tuberculosis
14
-
nicotinic acid hydrazide
-
Mycobacterium tuberculosis
14
-
benzoic acid hydrazide
-
Mycobacterium tuberculosis
20
-
4-hydroxybenzhydrazide
-
Mycobacterium tuberculosis
51
-
4-fluoroaniline
-
Mycobacterium tuberculosis
102
-
isoniazid
-
Mycobacterium tuberculosis

Localization

Localization Comment Organism GeneOntology No. Textmining
cytosol
-
Mycobacterium tuberculosis 5829
-

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
30000
-
SDS-PAGE Mycobacterium tuberculosis
30000
-
determined by SDS-PAGE Mycobacterium tuberculosis

Organism

Organism UniProt Comment Textmining
Mycobacterium tuberculosis
-
-
-

Purification (Commentary)

Purification (Comment) Organism
using a Ni-NTA and a Superdex S-75 column Mycobacterium tuberculosis
using Ni-NTA chromatography. Approximately 2.7 mg of purified enzyme is obtained per liter of culture Mycobacterium tuberculosis

Reaction

Reaction Comment Organism Reaction ID
acetyl-CoA + an arylamine = CoA + an N-acetylarylamine steady-state kinetic analysis of TBNAT reveals that the enzyme catalyzes the reaction via a bi-bi ping-pong kinetic mechanism: enzyme initially reacts with AcCoA to generate the first product, CoA, and the acetyl-enzyme intermediate (E-Ac). The amine binds and reacts with E-Ac to generate the second product, acetylated amine, and regenerate the free enzyme. The pH dependence of the kinetic parameters reveals that one enzyme group must be deprotonated for optimal catalytic activity and that two amino acid residues at the active site of the free enzyme are involved in binding and/or catalysis. Solvent kinetic isotope effects suggest that proton transfer steps are not rate-limiting in the overall reaction for substituted aniline substrates but become rate-limiting when poor hydrazide substrates are used Mycobacterium tuberculosis

Storage Stability

Storage Stability Organism
-20°C, 50% glycerol Mycobacterium tuberculosis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
acetyl-CoA + 2-amino-4-chlorophenol
-
Mycobacterium tuberculosis CoA + N-(5-chloro-2-hydroxyphenyl)acetamide
-
?
acetyl-CoA + 2-amino-4-chlorophenol
-
Mycobacterium tuberculosis CoA + N-acetyl-2-amino-4-chlorophenol
-
?
acetyl-CoA + 2-amino-4-methylphenol
-
Mycobacterium tuberculosis CoA + N-(2-hydroxy-5-methylphenyl)acetamide
-
?
acetyl-CoA + 2-amino-4-methylphenol
-
Mycobacterium tuberculosis CoA + N-acetyl-2-amino-4-methylphenol
-
?
acetyl-CoA + 2-aminophenol
-
Mycobacterium tuberculosis CoA + N-(2-hydroxyphenyl)acetamide
-
?
acetyl-CoA + 2-aminophenol
-
Mycobacterium tuberculosis CoA + N-acetyl-2-aminophenol
-
?
acetyl-CoA + 3-amino-4-hydroxybenzoic acid
-
Mycobacterium tuberculosis CoA + 3-(acetylamino)-4-hydroxybenzoic acid
-
?
acetyl-CoA + 3-amino-4-hydroxybenzoic acid
-
Mycobacterium tuberculosis CoA + N-acetyl-3-amino-4-hydroxybenzoic acid
-
?
acetyl-CoA + 4-amino-3-hydroxybenzoic acid
-
Mycobacterium tuberculosis CoA + 4-(acetylamino)-3-hydroxybenzoic acid
-
?
acetyl-CoA + 4-amino-3-hydroxybenzoic acid
-
Mycobacterium tuberculosis CoA + N-acetyl-4-amino-3-hydroxybenzoic acid
-
?
acetyl-CoA + 4-chloroaniline
-
Mycobacterium tuberculosis CoA + N-acetyl-4-chloroaniline
-
?
acetyl-CoA + 4-chloroaniline
-
Mycobacterium tuberculosis CoA + N-(4-chlorophenyl)acetamide
-
?
acetyl-CoA + 4-fluoroaniline
-
Mycobacterium tuberculosis CoA + N-(4-fluorophenyl)acetamide
-
?
acetyl-CoA + 4-fluoroaniline
-
Mycobacterium tuberculosis CoA + N-acetyl-4-fluoroaniline
-
?
acetyl-CoA + 4-hydroxybenzhydrazide
-
Mycobacterium tuberculosis CoA + N'-acetyl-4-hydroxybenzohydrazide
-
?
acetyl-CoA + 4-hydroxybenzhydrazide
-
Mycobacterium tuberculosis CoA + N-acetyl-4-hydroxybenzhydrazide
-
?
acetyl-CoA + 4-methylaniline
-
Mycobacterium tuberculosis CoA + N-acetyl-4-methylaniline
-
?
acetyl-CoA + 4-methylaniline
-
Mycobacterium tuberculosis CoA + N-(4-methylphenyl)acetamide
-
?
acetyl-CoA + aniline
-
Mycobacterium tuberculosis CoA + N-acetyl-aniline
-
?
acetyl-CoA + aniline
-
Mycobacterium tuberculosis CoA + N-phenylacetamide
-
?
acetyl-CoA + anisidine
-
Mycobacterium tuberculosis CoA + N-acetyl-anisidine
-
?
acetyl-CoA + anisidine
-
Mycobacterium tuberculosis CoA + N-(4-methoxyphenyl)acetamide
-
?
acetyl-CoA + benzoic acid hydrazide
-
Mycobacterium tuberculosis CoA + N'-acetylbenzohydrazide
-
?
acetyl-CoA + benzoic acid hydrazide
-
Mycobacterium tuberculosis CoA + N-acetyl-benzoic acid hydrazide
-
?
acetyl-CoA + hydralazine
-
Mycobacterium tuberculosis CoA + N'-phthalazin-1-ylacetohydrazide
-
?
acetyl-CoA + hydralazine
-
Mycobacterium tuberculosis CoA + N-acetyl-hydralazine
-
?
acetyl-CoA + isoniazid
-
Mycobacterium tuberculosis CoA + N-acetyl-isoniazid
-
?
acetyl-CoA + isoniazid
-
Mycobacterium tuberculosis CoA + N'-acetylpyridine-4-carbohydrazide
-
?
acetyl-CoA + nicotinic acid hydrazide
-
Mycobacterium tuberculosis CoA + N'-acetylpyridine-3-carbohydrazide
-
?
acetyl-CoA + nicotinic acid hydrazide
-
Mycobacterium tuberculosis CoA + N-acetyl-nicotinic acid hydrazide
-
?
acetyl-CoA + p-aminobenzoic acid
-
Mycobacterium tuberculosis CoA + N-acetyl-p-aminobenzoic acid
-
?
acetyl-CoA + p-aminobenzoic acid
-
Mycobacterium tuberculosis CoA + 4-(acetylamino)benzoic acid
-
?
acetyl-CoA + sulfamethoxazole
-
Mycobacterium tuberculosis CoA + N-[4-[(5-methylisoxazol-3-yl)sulfamoyl]phenyl]acetamide
-
?
additional information results confirm that substituted anilines are excellent substrates and that TBNAT is ineffective in hydrazide N-acetylation Mycobacterium tuberculosis ?
-
?

Synonyms

Synonyms Comment Organism
arylamine N-acetyltransferase
-
Mycobacterium tuberculosis
NAT
-
Mycobacterium tuberculosis
TBNAT
-
Mycobacterium tuberculosis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Mycobacterium tuberculosis
25
-
activity assay Mycobacterium tuberculosis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.2
-
4-chloroaniline
-
Mycobacterium tuberculosis
0.28
-
aniline
-
Mycobacterium tuberculosis
0.6
-
nicotinic acid hydrazide
-
Mycobacterium tuberculosis
0.7
-
p-Aminobenzoic acid
-
Mycobacterium tuberculosis
0.8
-
4-methylaniline
-
Mycobacterium tuberculosis
0.9
-
4-fluoroaniline
-
Mycobacterium tuberculosis
1.9
-
benzoic acid hydrazide
-
Mycobacterium tuberculosis
2.9
-
anisidine
-
Mycobacterium tuberculosis
4.9
-
isoniazid
-
Mycobacterium tuberculosis
10.9
-
4-hydroxybenzhydrazide
-
Mycobacterium tuberculosis
21
-
2-amino-4-chlorophenol
-
Mycobacterium tuberculosis
44
-
hydralazine
-
Mycobacterium tuberculosis
51
-
2-Aminophenol
-
Mycobacterium tuberculosis
66
-
4-amino-3-hydroxybenzoic acid
-
Mycobacterium tuberculosis
70
-
2-amino-4-methylphenol
-
Mycobacterium tuberculosis
94
-
3-Amino-4-hydroxybenzoic acid
-
Mycobacterium tuberculosis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.7
-
assay at Mycobacterium tuberculosis
7.7
-
activity assay Mycobacterium tuberculosis

Cofactor

Cofactor Comment Organism Structure
acetyl-CoA
-
Mycobacterium tuberculosis