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Literature summary extracted from

  • Bzymek, K.P.; Moulin, A.; Swierczek, S.I.; Ringe, D.; Petsko, G.A.; Bennett, B.; Holz, R.C.
    Kinetic, spectroscopic, and X-ray crystallographic characterization of the functional E151H aminopeptidase from Aeromonas proteolytica (2005), Biochemistry, 44, 12030-12040.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.4.11.10 expression of wild-type and mutant E151H enzymes in Escherichia coli strain BL21(DE3) Vibrio proteolyticus

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
3.4.11.10 purified recombinant E151H mutant holoenzyme, 15 mg/ml protein in 10 mM HEPES, pH 7.2, 10 mM KSCN, and 0.4 M NaCl,vapour diffusion mehtod, the precipitation solution contains 100 mM HEPES, pH 7.2, 100 mM KSCN, and 4.5 M NaCl, 2-3 days, X-ray diffraction structure determination and analysis at 1.9 A resolution and room temperature Vibrio proteolyticus

Protein Variants

EC Number Protein Variants Comment Organism
3.4.11.10 E151H site-directed mutagenesis, the mutant enzyme shows a highly reduced reaction rate compared to the wild-type enzyme Vibrio proteolyticus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.4.11.10 additional information
-
 additional information pH-dependence of kinetics, isothermal titration measurement, thermodynamics, overview Vibrio proteolyticus

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.4.11.10 Co2+ can substitute for Zn2+ Vibrio proteolyticus
3.4.11.10 additional information detailed metal binding structure analysis, overview, the first metal ion in the dinuclear metal center is in a hexacoordinate/pentacoordinate equilibrium, while the second metal ion is six-coordinate Vibrio proteolyticus
3.4.11.10 Zn2+ dinuclear Zn2+ metal center, the binding of Zn2+ to the E151H mutant is much more weak than to the wild-type enzyme Vibrio proteolyticus

Organism

EC Number Organism UniProt Comment Textmining
3.4.11.10 Vibrio proteolyticus Q01693
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.4.11.10 recombinant wild-type and mutant E151H enzymes from Escherichia coli strain BL21(DE3) by ammonium sulfate fractionation, dialysis, hydrophobic interaction and anion exchange chromatography Vibrio proteolyticus

Reaction

EC Number Reaction Comment Organism Reaction ID
3.4.11.10 Release of an N-terminal amino acid, preferentially leucine, but not glutamic or aspartic acids. active site residue Glu151 is essential for activity acting as a general acid/base during the catalytic reaction, mechanism of peptide hydrolysis Vibrio proteolyticus
a

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.4.11.10 L-Leu-4-nitroanilide + H2O
-
 Vibrio proteolyticus L-Leu + 4-nitroaniline
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.4.11.10 AAP
-
 Vibrio proteolyticus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.4.11.10 0.033
-
 L-Leu-4-nitroanilide pH 8.0, recombinant mutant E151H Vibrio proteolyticus
3.4.11.10 73
-
 L-Leu-4-nitroanilide pH 8.0, recombinant wild-type enzyme Vibrio proteolyticus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.4.11.10 7.5 8.5 assay at Vibrio proteolyticus

pH Range

EC Number pH Minimum pH Maximum Comment Organism
3.4.11.10 5.5 10.8 pH-profile of mutant E151H Vibrio proteolyticus