Literature summary for 3.4.11.10 extracted from

Kale, A.; Pijning, T.; Sonke, T.; Dijkstra, B.W.; Thunnissen, A.M.
Crystal structure of the leucine aminopeptidase from Pseudomonas putida reveals the molecular basis for its enantioselectivity and broad substrate specificity (2010), J. Mol. Biol., 398, 703-714.

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Application

Application	Comment	Organism
synthesis	LAP is an important enzyme for the industrial production of enantiomerically pure amino acids	Pseudomonas putida

Cloned(Commentary)

Cloned (Comment)	Organism
expression in Escherichia coli	Pseudomonas putida

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified enzyme in complex with inhibitor bestatin at pH 5.2 and pH 9.5, hanging drop vapour diffusion, 8 mg/ml protein in 20 mM HEPESKOH, pH 8.0, 1 mM DTT, is mixed with 11% w/v PEG 8000, 0.2 M sodium formate, 0.1 M MesNaOH, pH 5.2, 1 mM NaN3 at 5°C, or 4 mg/ml protein solution is mixed with 15% w/v PEG 1500, 0.1 M propionic acid, cacodylate, bis-Tris propane cocktail buffer, pH 9.5 at 23°C, three days, X-ray diffraction structure determination and analysis at 1.5-2.75 A resolution, modelling	Pseudomonas putida

Crystallization (Comment)

Organism

purified enzyme in complex with inhibitor bestatin at pH 5.2 and pH 9.5, hanging drop vapour diffusion, 8 mg/ml protein in 20 mM HEPESKOH, pH 8.0, 1 mM DTT, is mixed with 11% w/v PEG 8000, 0.2 M sodium formate, 0.1 M MesNaOH, pH 5.2, 1 mM NaN3 at 5°C, or 4 mg/ml protein solution is mixed with 15% w/v PEG 1500, 0.1 M propionic acid, cacodylate, bis-Tris propane cocktail buffer, pH 9.5 at 23°C, three days, X-ray diffraction structure determination and analysis at 1.5-2.75 A resolution, modelling

Pseudomonas putida

Inhibitors

Inhibitors	Comment	Organism	Structure
bestatin	binding structure and comparison, overview	Pseudomonas putida

Metals/Ions

Metals/Ions	Comment	Organism
Mn2+	at pH 9.5, the active site contains two metal ions, one identified as Mn2+ or Zn2+ at site 1, and the other as Zn2+ at site 2. Mn2+ has a significant activation effect when bound to site 1 of ppLAP	Pseudomonas putida
additional information	ppLAP requires the presence of divalent metal ions for its activity, in particular Zn2+ and/or Mn2+. At pH 5.2, the active site of ppLAP is highly disordered and metal ions are absent, most probably due to full protonation of one of the metal-interacting residues, Lys267	Pseudomonas putida
Zn2+	LAP is zinc-dependent. At pH 9.5, the active site contains two metal ions, one identified as Mn2+ or Zn2+ at site 1, and the other as Zn2+ at site 2	Pseudomonas putida

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
270000	-	recombinant enzyme, gel filtration	Pseudomonas putida

Organism

Organism	UniProt	Comment	Textmining
Pseudomonas putida	O86436	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant enzyme from Escherichia coli by cation exchange chromatography and gel filtration	Pseudomonas putida

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
additional information	LAP shows broad substrate specificity and high enantioselectivity, structure-function relationship, active site metal composition and metal-dependent activity, overview	Pseudomonas putida	?	-	?

Synonyms

Synonyms	Comment	Organism
LAP	-	Pseudomonas putida
leucine aminopeptidase	-	Pseudomonas putida

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8	-	assay at	Pseudomonas putida

pH Range

pH Minimum	pH Maximum	Comment	Organism
additional information	-	the enzyme is inactive at low pH	Pseudomonas putida