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Literature summary for 3.4.11.10 extracted from

  • Acosta, D.; Cancela, M.; Piacenza, L.; Roche, L.; Carmona, C.; Tort, J.F.
    Fasciola hepatica leucine aminopeptidase, a promising candidate for vaccination against ruminant fasciolosis (2008), Mol. Biochem. Parasitol., 158, 52-64.
    View publication on PubMed
Search for more literature for 3.4.11.10

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli TOP 10 cells Fasciola hepatica

Inhibitors

Inhibitors Comment Organism Structure
1,10-phenanthroline the recombinant enzyme shows 18.8% relative activity in the presence of 1 mM 1,10-phenanthroline on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica
Aprotinin the recombinant enzyme shows 58.9% relative activity in the presence of 0.001 mM aprotinin on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica
bestatin strongly inhibited by bestatin, 20.5% relative activity in the presence of bestatin Fasciola hepatica
Ca2+ 44% relative activity at 10 mM Ca2+ Fasciola hepatica
dithiothreitol the recombinant enzyme shows 20.5% relative activity in the presence of 5 mM dithiothreitol on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica
E-64 the recombinant enzyme shows 43% relative activity in the presence of 0.005 mM E-64 on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica
L-cysteine strongly inhibited by cysteine Fasciola hepatica
leupeptin the recombinant enzyme shows 32.7% relative activity in the presence of 0.01 mM leupeptin on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica
N-alpha-tosyl-L-lysine chlormethyl ketone the recombinant enzyme shows 28.9% relative activity in the presence of 0.005 mM N-alpha-tosyl-L-lysine chlormethyl ketone on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica
pepstatin A the recombinant enzyme shows 26.1% relative activity in the presence of 1 mM pepstatin A on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica
phenylmethylsulfonyl fluoride the recombinant enzyme shows 24.5% relative activity in the presence of 1 mM phenylmethylsulfonyl fluoride on hydrolysis of L-Leu-7-amido-4-methylcoumarin Fasciola hepatica

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.038
-
 L-Leu-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1 mM MnCl2, at 37°C Fasciola hepatica
0.06
-
 L-Thr-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1 mM MnCl2, at 37°C Fasciola hepatica
0.066
-
 L-Phe-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1 mM MnCl2, at 37°C Fasciola hepatica
0.077
-
 L-Cys-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1 mM MnCl2, at 37°C Fasciola hepatica
0.078
-
 L-Arg-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1 mM MnCl2, at 37°C Fasciola hepatica
0.093
-
 L-Met-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1 mM MnCl2, at 37°C Fasciola hepatica
0.231
-
 L-Ala-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1 mM MnCl2, at 37°C Fasciola hepatica

Localization

Localization Comment Organism GeneOntology No. Textmining
cytosol
-
 Fasciola hepatica 5829
-

Metals/Ions

Metals/Ions Comment Organism Structure
Co2+ CoCl2 at 1mM increases the specific activity 18fold Fasciola hepatica
Mg2+ MgCl2 shows an optimal concentration at 10 mM that increases specific activity by 12fold Fasciola hepatica
Mn2+ recombinant LAP activity is progressively activated by MnCl2 (0.001-10 mM) with an optimum at 1 mM, yielding a 43fold increase Fasciola hepatica
additional information no relevant stimulatory effect with NiCl2, FeSO4 and CuSO4 Fasciola hepatica
Zn2+ ZnCl2 modestly affects the enzymatic activity Fasciola hepatica

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
67000
-
 recombinant enzyme, SDS-PAGE Fasciola hepatica

Organism

Organism UniProt Comment Textmining
Fasciola hepatica
-
 LAP preferentially localizes inside the epithelial cells that line the alimentary tract of the adult worm
-

Purification (Commentary)

Purification (Comment) Organism
native and recombinant enzymes are purified by Ni-NTA column chromatography, Superdex 200 gel filtration and bestatin-affinity chromatography Fasciola hepatica

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.0001
-
 recombinant enzyme, using L-Glu-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0002
-
 native enzyme, using L-Glu-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0003
-
 native enzyme, using L-Asp-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0005
-
 native enzyme, using Gly-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0006
-
 native enzyme, using L-Val-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0012
-
 recombinant enzyme, using L-Asp-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0019
-
 native enzyme, using L-Ser-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0019
-
 recombinant enzyme, using Gly-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0021
-
 recombinant enzyme, using L-Ser-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0025
-
 recombinant enzyme, using L-Val-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0032
-
 native enzyme, using L-Pro-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0054
-
 native enzyme, using L-Tyr-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0056
-
 recombinant enzyme, using L-Pro-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0062
-
 recombinant enzyme, using L-Ile-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0064
-
 recombinant enzyme, using L-Tyr-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0068
-
 recombinant enzyme, using L-Phe-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0075
-
 recombinant enzyme, using L-Ala-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0083
-
 native enzyme, using L-Ala-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0093
-
 native enzyme, using L-Ile-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0141
-
 native enzyme, using L-Phe-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0253
-
 recombinant enzyme, using L-Met-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0291
-
 recombinant enzyme, using L-Arg-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0355
-
 recombinant enzyme, using L-Thr-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0657
-
 native enzyme, using L-Met-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0694
-
 native enzyme, using L-Thr-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0752
-
 native enzyme, using L-Arg-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0831
-
 recombinant enzyme, using L-Cys-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.0953
-
 native enzyme, using L-Cys-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.1344
-
 native enzyme, using L-Leu-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica
0.2145
-
 recombinant enzyme, using L-Leu-7-amido-4-methylcoumarin as substrate, at 37°C Fasciola hepatica

Storage Stability

Storage Stability Organism
-80°C, 3 months, no appreciable loss of activity Fasciola hepatica

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
Gly-7-amido-4-methylcoumarin + H2O very weak activity Fasciola hepatica Gly + 7-amino-4-methylcoumarin
-
 ?
L-Ala-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Ala + 7-amino-4-methylcoumarin
-
 ?
L-Arg-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Arg + 7-amino-4-methylcoumarin
-
 ?
L-Asp-7-amido-4-methylcoumarin + H2O very weak activity Fasciola hepatica L-Asp + 7-amino-4-methylcoumarin
-
 ?
L-Cys-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Cys + 7-amino-4-methylcoumarin
-
 ?
L-Glu-7-amido-4-methylcoumarin + H2O very weak activity Fasciola hepatica L-Glu + 7-amino-4-methylcoumarin
-
 ?
L-Ile-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Ile + 7-amino-4-methylcoumarin
-
 ?
L-Leu-7-amido-4-methylcoumarin + H2O preferred substrate Fasciola hepatica L-Leu + 7-amino-4-methylcoumarin
-
 ?
L-Met-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Met + 7-amino-4-methylcoumarin
-
 ?
L-Phe-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Phe + 7-amino-4-methylcoumarin
-
 ?
L-Pro-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Pro + 7-amino-4-methylcoumarin
-
 ?
L-Ser-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Ser + 7-amino-4-methylcoumarin
-
 ?
L-Thr-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Thr + 7-amino-4-methylcoumarin
-
 ?
L-Tyr-7-amido-4-methylcoumarin + H2O
-
 Fasciola hepatica L-Tyr + 7-amino-4-methylcoumarin
-
 ?
L-Val-7-amido-4-methylcoumarin + H2O very weak activity Fasciola hepatica L-Val + 7-amino-4-methylcoumarin
-
 ?

Synonyms

Synonyms Comment Organism
LAP
-
 Fasciola hepatica
leucine aminopeptidase
-
 Fasciola hepatica
Leucyl aminopeptidase
-
 Fasciola hepatica

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 the recombinant enzyme is active at a wide range of temperatures, with two peaks, at 37 and 50°C Fasciola hepatica
50
-
 the recombinant enzyme is active at a wide range of temperatures, with two peaks, at 37 and 50°C Fasciola hepatica

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
50 60 preheating of recombinant LAP at 50°C produces the maximum specific activity while at higher temperatures (above 60°C) the enzyme results inactivated Fasciola hepatica

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
16200
-
 L-Thr-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1mM MnCl2, at 37°C Fasciola hepatica
33800
-
 L-Phe-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1mM MnCl2, at 37°C Fasciola hepatica
59900
-
 L-Arg-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1mM MnCl2, at 37°C Fasciola hepatica
192000
-
 L-Ala-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1mM MnCl2, at 37°C Fasciola hepatica
246000
-
 L-Leu-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1mM MnCl2, at 37°C Fasciola hepatica
283200
-
 L-Met-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1mM MnCl2, at 37°C Fasciola hepatica
288100
-
 L-Cys-7-amido-4-methylcoumarin recombinant enzyme, in 0.1 M glycine buffer, pH 8.5 containing 1mM MnCl2, at 37°C Fasciola hepatica

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8 8.5
-
 Fasciola hepatica

pH Range

pH Minimum pH Maximum Comment Organism
7 9 optimal activity is observed at pH 8-8.5, exhibiting a rapid decline under mildly acidic (below pH 7) and basic (above pH 9) conditions Fasciola hepatica

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.000001
-
 bestatin recombinant enzyme, at 37°C Fasciola hepatica
0.000025
-
 L-cysteine recombinant enzyme, at 37°C Fasciola hepatica