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Literature summary for 3.5.1.81 extracted from

  • Yano, S.; Haruta, H.; Ikeda, T.; Kikuchi, T.; Murakami, M.; Moriguchi, M.; Wakayama, M.
    Engineering the substrate specificity of Alcaligenes D-aminoacylase useful for the production of D-amino acids by optical resolution (2011), J. Chromatogr. B, 879, 3247-3252.
    View publication on PubMed
Search for more literature for 3.5.1.81

Application

Application Comment Organism
synthesis the F191W mutant is considered to be useful for the enzymatic production of D-Trp which is an important building block of some therapeutic drugs Achromobacter xylosoxidans

Cloned(Commentary)

Cloned (Comment) Organism
wild type and mutant enzymes are expressed in Escherichia coli JM109 cells Achromobacter xylosoxidans

Protein Variants

Protein Variants Comment Organism
F191W the catalytic efficiency of the mutant toward N-acetyl-D-Trp and N-acetyl-D-Ala is enhanced by 15.6 and 1.5folds, respectively, compared with that of the wild type enzyme. The mutant retains its catalytic efficiency toward N-acetyl-D-Met compared with that of the wild type enzyme Achromobacter xylosoxidans
L298A the catalytic efficiency of the mutant toward N-acetyl-D-Trp is enhanced by 4.4folds compared with that of the wild type enzyme. The catalytic efficiency for N-acetyl-D-Met and N-acetyl-D-Ala decrease to between 2.5 and 5% of that of the wild type enzyme, respectively Achromobacter xylosoxidans
L298F the mutant shows rather lower activities toward all substrates tested including N-acetyl-D-Met compared with the wild type enzyme Achromobacter xylosoxidans
M346F the mutant shows rather lower activities toward all substrates tested including N-acetyl-D-Met compared with the wild type enzyme Achromobacter xylosoxidans
V297F the mutant shows partially lower activities toward all substrates tested compared with the wild type enzyme Achromobacter xylosoxidans

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.03
-
 N-Acetyl-D-Met mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
2.97
-
 N-Acetyl-D-Trp mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
4.8
-
 N-Acetyl-D-Met wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
6.23
-
 N-Acetyl-D-Trp wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
6.36
-
 N-Acetyl-D-Trp mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
38.61
-
 N-Acetyl-D-Ala mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
41
-
 N-Acetyl-D-Ala wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
56.4
-
 N-Acetyl-D-Met mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
330
-
 N-Acetyl-D-Ala mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
52000
-
 x * 52000, SDS-PAGE Achromobacter xylosoxidans

Organism

Organism UniProt Comment Textmining
Achromobacter xylosoxidans
-
 subsp. xylosoxydans
-
Achromobacter xylosoxidans A-6
-
 subsp. xylosoxydans
-

Purification (Commentary)

Purification (Comment) Organism
DEAE-Toyopearl column chromatography and butyl-Cellulofine column chromatography Achromobacter xylosoxidans

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
N-Acetyl-D-Ala + H2O
-
 Achromobacter xylosoxidans acetate + D-Ala
-
 ?
N-Acetyl-D-Ala + H2O
-
 Achromobacter xylosoxidans A-6 acetate + D-Ala
-
 ?
N-Acetyl-D-Met + H2O preferred substrate Achromobacter xylosoxidans acetate + D-Met
-
 ?
N-Acetyl-D-Met + H2O preferred substrate Achromobacter xylosoxidans A-6 acetate + D-Met
-
 ?
N-Acetyl-D-Trp + H2O poor substrate Achromobacter xylosoxidans acetate + D-Trp
-
 ?
N-Acetyl-D-Trp + H2O poor substrate Achromobacter xylosoxidans A-6 acetate + D-Trp
-
 ?

Subunits

Subunits Comment Organism
? x * 52000, SDS-PAGE Achromobacter xylosoxidans

Synonyms

Synonyms Comment Organism
D-Aminoacylase
-
 Achromobacter xylosoxidans
D-NAase
-
 Achromobacter xylosoxidans

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
5.97
-
 N-Acetyl-D-Ala mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
89.5
-
 N-Acetyl-D-Trp wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
229
-
 N-Acetyl-D-Ala wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
330
-
 N-Acetyl-D-Ala mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
404
-
 N-Acetyl-D-Trp mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
463
-
 N-Acetyl-D-Met mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
669
-
 N-Acetyl-D-Trp mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
829
-
 N-Acetyl-D-Met mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
1826
-
 N-Acetyl-D-Met wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.02
-
 N-Acetyl-D-Ala mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
5.58
-
 N-Acetyl-D-Ala wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
8.21
-
 N-Acetyl-D-Met mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
8.56
-
 N-Acetyl-D-Ala mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
14.4
-
 N-Acetyl-D-Trp wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
63.5
-
 N-Acetyl-D-Trp mutant enzyme L298A, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
225
-
 N-Acetyl-D-Trp mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
381
-
 N-Acetyl-D-Met wild type enzyme, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans
407
-
 N-Acetyl-D-Met mutant enzyme F191W, in 100 mM HEPES (pH 7.0), at 30°C Achromobacter xylosoxidans