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Literature summary extracted from

  • Rueda, D.; Bernard, C.; Gandy, L.; Capton, E.; Boudjelloul, R.; Brossier, F.; Veziris, N.; Zimic, M.; Sougakoff, W.
    Estimation of pyrazinamidase activity using a cell-free in vitro synthesis of pnca and its association with pyrazinamide susceptibility in Mycobacterium tuberculosis (2018), Int. J. Mycobacteriol., 7, 16-25 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.1.B15 gene pncA, PCR-based in vitro expression, and recombinant expression of His-tagged mutant enzyme in Escherichia coli strain BL21(DE3), subcloning in Escherichia coli strain TOP10 Mycobacterium tuberculosis variant bovis
3.5.1.B15 gene pncA, PCR-based in vitro expression, and recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3), subcloning in Escherichia coli strain TOP10 Mycobacterium tuberculosis

Protein Variants

EC Number Protein Variants Comment Organism
3.5.1.B15 A146V naturally occuring mutation, the mutant shows 72% reduced activity compared to wild-type, resistant strain Mycobacterium tuberculosis
3.5.1.B15 D63A naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 G97D naturally occuring mutation, the mutant shows 90% reduced activity compared to wild-type, resistant strain Mycobacterium tuberculosis
3.5.1.B15 H51P naturally occuring mutation, inactive mutant, resistant strain Mycobacterium tuberculosis
3.5.1.B15 H51R naturally occuring mutation, inactive mutant, resistant strain Mycobacterium tuberculosis
3.5.1.B15 H57D naturally occuring mutation, inactive mutant, resistant strain Mycobacterium tuberculosis
3.5.1.B15 H57D site-directed mutagenesis, inactive mutant Mycobacterium tuberculosis variant bovis
3.5.1.B15 I5T naturally occuring mutation, the mutant shows 88% reduced activity compared to wild-type, resistant strain Mycobacterium tuberculosis
3.5.1.B15 I6L naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 L85R naturally occuring mutation, inactive mutant, resistant strain Mycobacterium tuberculosis
3.5.1.B15 M175V naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 additional information analysis of the specific pyrazinamidase (PZase) activity of PncA mutants via PCR-based in vitro-synthesized-PZase assay and correlation of the results to the pyrazinamide (PZA) susceptibility phenotype determined by culture in acidic agar medium at pH 6.0. A set of 23 clinical isolates displaying mutated pncA genes (11 PZA-resistant and 12 PZA-susceptible) and 55 PZA-susceptible clinical strains displaying a wild-type pncA gene are tested. Among the 23 mutants tested, 4 corresponded to mutations not reported before (I5T, Y99S, T142R, and P77L/V131G). Of the 11 PncA mutants expressed from PZA-resistant clinical isolates, 9 are expressed in vitro at yields above 50% relative to the wild-type enzyme. Among them, 6 enzymes (T47P, H51P, H51R, H57D, L85R and T142R) show no detectable activity, while the relative activities for the 3 others, V9A, G97D, and A146V, are low compared to the wild-type PZase. The remaining two mutants, I5T and V9G, presented very low relative expression (5%) and relative activities values of 12 and 1%, respectively. Twelve mutants are expressed from PZA-susceptible isolates. Their expression arte is similar to the wild-type enzyme, and they behave as active pyrazinamidase with specific relative activities ranging from 34 to 314%. Finally, discrepant results are observed for two mutants, V7A and P62T Mycobacterium tuberculosis
3.5.1.B15 P62T naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 P77L/V131G naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 R140S naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 T142R naturally occuring mutation, resistant strain Mycobacterium tuberculosis
3.5.1.B15 T142R naturally occuring mutation, inactive mutant, resistant strain Mycobacterium tuberculosis
3.5.1.B15 T167I naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 T47A naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 T47P naturally occuring mutation, inactive mutant, resistant strain Mycobacterium tuberculosis
3.5.1.B15 T87M site-directed mutagenesis, active mutant, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 V7A naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 V9A naturally occuring mutation, the mutant shows 73% reduced activity compared to wild-type, resistant strain Mycobacterium tuberculosis
3.5.1.B15 V9G naturally occuring mutation, the mutant shows 99% reduced activity compared to wild-type, resistant strain Mycobacterium tuberculosis
3.5.1.B15 Y64D naturally occuring mutation, susceptible strain Mycobacterium tuberculosis
3.5.1.B15 Y99S naturally occuring mutation, susceptible strain Mycobacterium tuberculosis

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.5.1.B15 Fe2+ required, enzyme-bound Mycobacterium tuberculosis

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
3.5.1.B15 pyrazinamide + H2O Mycobacterium tuberculosis
-
 pyrazinoic acid + NH3
-
 ?
3.5.1.B15 pyrazinamide + H2O Mycobacterium tuberculosis variant bovis
-
 pyrazinoic acid + NH3
-
 ?
3.5.1.B15 pyrazinamide + H2O Mycobacterium tuberculosis H37Rv
-
 pyrazinoic acid + NH3
-
 ?
3.5.1.B15 pyrazinamide + H2O Mycobacterium tuberculosis ATCC 25618
-
 pyrazinoic acid + NH3
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
3.5.1.B15 Mycobacterium tuberculosis I6XD65
-
-
3.5.1.B15 Mycobacterium tuberculosis ATCC 25618 I6XD65
-
-
3.5.1.B15 Mycobacterium tuberculosis H37Rv I6XD65
-
-
3.5.1.B15 Mycobacterium tuberculosis variant bovis
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.1.B15 recombinant His-tagged mutant enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and ultrafiltration Mycobacterium tuberculosis variant bovis
3.5.1.B15 recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and ultrafiltration Mycobacterium tuberculosis

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.1.B15 additional information colorimetric assay using ferrous ammonium sulfate 20% and glycine-HCl Mycobacterium tuberculosis ?
-
 ?
3.5.1.B15 additional information colorimetric assay using ferrous ammonium sulfate 20% and glycine-HCl Mycobacterium tuberculosis variant bovis ?
-
 ?
3.5.1.B15 additional information colorimetric assay using ferrous ammonium sulfate 20% and glycine-HCl Mycobacterium tuberculosis H37Rv ?
-
 ?
3.5.1.B15 additional information colorimetric assay using ferrous ammonium sulfate 20% and glycine-HCl Mycobacterium tuberculosis ATCC 25618 ?
-
 ?
3.5.1.B15 pyrazinamide + H2O
-
 Mycobacterium tuberculosis pyrazinoic acid + NH3
-
 ?
3.5.1.B15 pyrazinamide + H2O
-
 Mycobacterium tuberculosis variant bovis pyrazinoic acid + NH3
-
 ?
3.5.1.B15 pyrazinamide + H2O
-
 Mycobacterium tuberculosis H37Rv pyrazinoic acid + NH3
-
 ?
3.5.1.B15 pyrazinamide + H2O
-
 Mycobacterium tuberculosis ATCC 25618 pyrazinoic acid + NH3
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.5.1.B15 additional information cf. EC 3.5.1.19 Mycobacterium tuberculosis
3.5.1.B15 additional information cf. EC 3.5.1.19 Mycobacterium tuberculosis variant bovis
3.5.1.B15 PncA
-
 Mycobacterium tuberculosis
3.5.1.B15 PncA
-
 Mycobacterium tuberculosis variant bovis
3.5.1.B15 pyrazinamidase/nicotinamidase
-
 Mycobacterium tuberculosis
3.5.1.B15 pyrazinamidase/nicotinamidase
-
 Mycobacterium tuberculosis variant bovis
3.5.1.B15 PZAse
-
 Mycobacterium tuberculosis
3.5.1.B15 PZAse
-
 Mycobacterium tuberculosis variant bovis

General Information

EC Number General Information Comment Organism
3.5.1.B15 malfunction estimation of pyrazinamidase activity using a cell-free in vitro synthesis of pncA and its association with pyrazinamide susceptibility in Mycobacterium tuberculosis, mutant phenotypes, overview Mycobacterium tuberculosis
3.5.1.B15 additional information catalytic Cys138 Mycobacterium tuberculosis