Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli TOP 10 cells | Fasciola hepatica |
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 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 | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
cytosol | - |
Fasciola hepatica | 5829 | - |
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 Maximum [Da] | Comment | Organism |
---|---|---|---|
67000 | - |
recombinant enzyme, SDS-PAGE | Fasciola hepatica |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Fasciola hepatica | - |
LAP preferentially localizes inside the epithelial cells that line the alimentary tract of the adult worm | - |
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 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 | Organism |
---|---|
-80°C, 3 months, no appreciable loss of activity | Fasciola hepatica |
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 | Comment | Organism |
---|---|---|
LAP | - |
Fasciola hepatica |
leucine aminopeptidase | - |
Fasciola hepatica |
Leucyl aminopeptidase | - |
Fasciola hepatica |
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 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 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 Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | 8.5 | - |
Fasciola hepatica |
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 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 |