Cloned (Comment) | Organism |
---|---|
gene LAP, sequence comparions and phylogenetic analysis, recombinant expression of His6-tagged TF-fused AcLAPr in Escherichia coli strain BL21, recombinant expression of EGFP-tagged enzyme in Acanthamoeba castellanii trophozoites | Acanthamoeba castellanii |
Protein Variants | Comment | Organism |
---|---|---|
additional information | AcLAP knockout using siRNA, AcLAP knockdown affects encystation of Acanthamoeba castellanii | Acanthamoeba castellanii |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
1,10-phenanthroline | abolishes enzyme activity at 10 mM | Acanthamoeba castellanii | |
bestatin | inhibits enzyme activity by 77% at 0.1 mM | Acanthamoeba castellanii | |
Co2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii | |
EDTA | abolishes enzyme activity at 1 mM | Acanthamoeba castellanii | |
Mn2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii | |
Ni2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii | |
Zn2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0038 | - |
L-methionyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii | |
0.0135 | - |
L-leucyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii | |
0.0414 | - |
L-arginyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
cytoplasm | AcLAP is mainly localized in the cytoplasm of Acanthamoeba castellanii | Acanthamoeba castellanii | 5737 | - |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Co2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii | |
Mg2+ | activates at 0.1-10 mM | Acanthamoeba castellanii | |
Mn2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii | |
additional information | AcLAPr activity is progressively enhanced by Ni2+, Mn2+, Mg2+, Zn2+, and Co2+ but not by Ca2+. Among the divalent metal ions tested, Ni2+ (0.1-1.0 mM) increases AcLAPr activity the most by 5.5fold. Higher concentrations of all metal ions of 10 mM, except of Mg2+, inhibit AcLAPr activity | Acanthamoeba castellanii | |
Ni2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii | |
Zn2+ | activates at 0.1-1.0 mM, inhibitory at 1.0-10 mM | Acanthamoeba castellanii |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Acanthamoeba castellanii | A0A060A630 | - |
- |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
cyst | AcLAP is highly expressed at a late stage of encystation, an approximate twofold increase is observed at 72 h post-encystation | Acanthamoeba castellanii | - |
trophozoite | the expression of AcLAP in trophozoites shows no significant change of its level during 24 h and 48 h | Acanthamoeba castellanii | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-arginyl-7-amido-4-methylcoumarin + H2O | low activity | Acanthamoeba castellanii | L-arginine + 7-amino-4-methylcoumarin | - |
? | |
L-leucyl-7-amido-4-methylcoumarin + H2O | moderate activity | Acanthamoeba castellanii | L-leucine + 7-amino-4-methylcoumarin | - |
? | |
L-methionyl-7-amido-4-methylcoumarin + H2O | high activity | Acanthamoeba castellanii | L-methionine + 7-amino-4-methylcoumarin | - |
? |
Subunits | Comment | Organism |
---|---|---|
? | x * 61800, about, sequence calculation | Acanthamoeba castellanii |
Synonyms | Comment | Organism |
---|---|---|
AcLAP | - |
Acanthamoeba castellanii |
M17 leucine aminopeptidase | - |
Acanthamoeba castellanii |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Acanthamoeba castellanii |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.13 | - |
L-arginyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii | |
0.33 | - |
L-methionyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii | |
0.83 | - |
L-leucyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Acanthamoeba castellanii |
General Information | Comment | Organism |
---|---|---|
evolution | phylogenetic analysis reveals that AcLAP clusters into a clade closely related with LAPs from Amoebozoa, including Dictyostelium sp.. The enzyme contains the highly conserved M17 aminopeptidase family signature sequence (NTDAEGRL, residues 425-432) and highly conserved amino acid residues for metal binding (D350, D368, D427, E429) and catalytic site formation (K357, R431). The enzyme belongs to the M17 family of proteases | Acanthamoeba castellanii |
malfunction | lack or downregulation of AcLAP activity causes ultrastructural changes of the cell wall in Acanthamoeba castellanii that are closely associated with inhibition of cyst formation | Acanthamoeba castellanii |
physiological function | AcLAP is a typical M17 family enzyme that plays an essential role during encystation of Acanthamoeba castellanii | Acanthamoeba castellanii |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
3.14 | - |
L-arginyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii | |
61.5 | - |
L-leucyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii | |
86.8 | - |
L-methionyl-7-amido-4-methylcoumarin | pH 8.0, 37°C, recombinant enzyme | Acanthamoeba castellanii |