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Literature summary extracted from

  • Steele, M.; Marcone, M.; Gyles, C.; Chan, V.L.; Odumeru, J.
    Enzymatic activity of Campylobacter jejuni hippurate hydrolase (2006), Protein Eng. Des. Sel., 19, 17-25.
    View publication on PubMed

Activating Compound

EC Number Activating Compound Comment Organism Structure
3.5.1.32 EDTA 23.9% activation at 10 mM Campylobacter jejuni

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.1.32 gene hipO, expression of His6-tagged enzyme in Escherichia coli strain M15 Campylobacter jejuni

Protein Variants

EC Number Protein Variants Comment Organism
3.5.1.32 D104N site-directed mutagenesis, inactive mutant Campylobacter jejuni
3.5.1.32 D76N site-directed mutagenesis, the mutant shows over 12fold reduced activity compared to the wild-type enzyme Campylobacter jejuni
3.5.1.32 E134Q site-directed mutagenesis, inactive mutant Campylobacter jejuni
3.5.1.32 E135Q site-directed mutagenesis, inactive mutant Campylobacter jejuni
3.5.1.32 H161A site-directed mutagenesis, inactive mutant Campylobacter jejuni
3.5.1.32 H356A site-directed mutagenesis, inactive mutant Campylobacter jejuni

Inhibitors

EC Number Inhibitors Comment Organism Structure
3.5.1.32 1,10-phenanthroline 99% inhibition at 0.5 mM Campylobacter jejuni
3.5.1.32 2-mercaptoethanol 20% inhibition at 2 mM Campylobacter jejuni
3.5.1.32 Ag+ 98% inhibition at 2 mM Campylobacter jejuni
3.5.1.32 Ca2+ 55% inhibition at 2 mM Campylobacter jejuni
3.5.1.32 Cd2+ 65% inhibition at 2 mM Campylobacter jejuni
3.5.1.32 Co2+ 70% inhibition at 2 mM Campylobacter jejuni
3.5.1.32 Cu2+ 78% inhibition at 2 mM Campylobacter jejuni
3.5.1.32 Fe2+
-
 Campylobacter jejuni
3.5.1.32 additional information no significant inhibition by E64, PMSF, and pepstatin at 0.5 mM Campylobacter jejuni
3.5.1.32 Ni2+ 65% inhibition at 2 mM Campylobacter jejuni
3.5.1.32 Zn2+ 35% inhibition at 2 mM Campylobacter jejuni

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.5.1.32 0.159
-
 N-benzoylglutamate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 0.169
-
 N-benzoylalanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 0.195
-
 N-benzoylphenylalanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 0.303
-
 N-benzoylmethionine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 0.594
-
 N-benzoylvaline pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 0.65
-
 hippurate pH 7.5, 37°C, native enzyme in cell extract Campylobacter jejuni
3.5.1.32 0.764
-
 hippurate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 1.769
-
 4-aminobenzoyl-glutamate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 2.778
-
 hippurate pH 7.5, 37°C, recombinant mutant D76N Campylobacter jejuni
3.5.1.32 3.377
-
 N-acetylglycine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 5.45
-
 N-acetyl-L-alanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.5.1.32 Ba2+ 10% activation at 10 mM Campylobacter jejuni
3.5.1.32 K+ 20% activation at 10 mM Campylobacter jejuni
3.5.1.32 Mg2+ 20% activation at 10 mM Campylobacter jejuni
3.5.1.32 Zn2+ 1.98 molecules of zinc per enzyme subunit molecule, metalloenzyme Campylobacter jejuni

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
3.5.1.32 42400
-
 4 * 42400, recombinant His6-tagged enzyme, SDS-PAGE Campylobacter jejuni
3.5.1.32 193000
-
 recombinant His6-tagged enzyme, gel filtration Campylobacter jejuni

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
3.5.1.32 hippurate + H2O Campylobacter jejuni
-
 benzoate + glycine
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
3.5.1.32 Campylobacter jejuni
-
 strain ATCC 43431, gene hipO
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.1.32 recombinant His6-tagged enzyme from Escherichia coli strain M15 by nickel affinity chromatography Campylobacter jejuni

Reaction

EC Number Reaction Comment Organism Reaction ID
3.5.1.32 hippurate + H2O = benzoate + glycine residues Asp76, Aps104, Glu134, Glu135, His161 and His356 are involved in zinc-binding and enzyme catalysis Campylobacter jejuni
a

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.5.1.32 120
-
 purified recombinant wild-type enzyme Campylobacter jejuni

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.1.32 4-aminobenzoyl-glutamate + H2O
-
 Campylobacter jejuni 4-aminobenzoate + glutamate
-
 ?
3.5.1.32 hippurate + H2O
-
 Campylobacter jejuni benzoate + glycine
-
 ?
3.5.1.32 hippurate + H2O i.e. N-benzoylglycine Campylobacter jejuni benzoate + glycine
-
 ?
3.5.1.32 additional information substrate specificity, the enzyme also shows metallocarboxypeptidase activity with carboxypeptidase substrates in vitro, displaying its highest activity against N-benzoyl-linked small aliphatic acids, overview, the enzyme does not hydrolyze typical substrates for aminopeptidases, lysine-p-nitroanilide, leucine-p-nitroanilide, or endopeptidases, N-benzoylarginine-p-nitroanilide, succinylphenylalanine-4-nitroanilide Campylobacter jejuni ?
-
 ?
3.5.1.32 N-acetylalanine + H2O
-
 Campylobacter jejuni acetate + alanine
-
 ?
3.5.1.32 N-acetylglycine + H2O
-
 Campylobacter jejuni acetate + glycine
-
 ?
3.5.1.32 N-benzoylalanine + H2O
-
 Campylobacter jejuni benzoate + alanine
-
 ?
3.5.1.32 N-benzoylglutamate + H2O
-
 Campylobacter jejuni benzoate + glutamate
-
 ?
3.5.1.32 N-benzoylmethionine + H2O
-
 Campylobacter jejuni benzoate + methionine
-
 ?
3.5.1.32 N-benzoylphenylalanine + H2O
-
 Campylobacter jejuni benzoate + phenylalanine
-
 ?
3.5.1.32 N-benzoylvaline + H2O
-
 Campylobacter jejuni benzoate + valine
-
 ?

Subunits

EC Number Subunits Comment Organism
3.5.1.32 More the enzyme structure shows highly ordered alpha-helix and beta-sheet sequences Campylobacter jejuni
3.5.1.32 tetramer 4 * 42400, recombinant His6-tagged enzyme, SDS-PAGE Campylobacter jejuni

Synonyms

EC Number Synonyms Comment Organism
3.5.1.32 hippurate hydrolase
-
 Campylobacter jejuni

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.5.1.32 50
-
-
 Campylobacter jejuni

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.5.1.32 0.052 2.1 N-acetylglycine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 0.17
-
 N-benzoylglutamate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 0.5
-
 N-benzoylvaline pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 1.4
-
 hippurate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 2 8 N-acetyl-L-alanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 2.32
-
 hippurate pH 7.5, 37°C, recombinant mutant D76N Campylobacter jejuni
3.5.1.32 2.6
-
 N-benzoylmethionine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 5.95
-
 N-benzoylphenylalanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 10.18
-
 N-acetylglycine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 13.33
-
 N-benzoylvaline pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 14.75
-
 4-aminobenzoyl-glutamate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 22.12
-
 N-acetyl-L-alanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 29.38
-
 N-benzoylalanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 48.72
-
 N-benzoylglutamate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 57.13
-
 N-benzoylmethionine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 81
-
 N-benzoylalanine pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni
3.5.1.32 98.73
-
 hippurate pH 7.5, 37°C, purified recombinant wild-type enzyme Campylobacter jejuni

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.5.1.32 7.5
-
-
 Campylobacter jejuni