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Literature summary extracted from

  • Honda, Y.; Arai, S.; Suzuki, K.; Kitaoka, M.; Fushinobu, S.
    The crystal structure of an inverting glycoside hydrolase family 9 exo-beta-D-glucosaminidase and the design of glycosynthase (2016), Biochem. J., 473, 463-472 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.2.1.165 wild type and mutant enzymes are expressed in Escherichia coli BL21(DE3) cells Photobacterium profundum

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
3.2.1.165 sitting drop vapor diffusion method, using 10% (w/v) PEG 6000, 0.1 M HEPES/NaOH (pH 7.5) and 5% (w/v) 2-methyl-2,4-pentanediol Photobacterium profundum

Protein Variants

EC Number Protein Variants Comment Organism
3.2.1.165 D139A the mutation significantly decreases the hydrolytic activity compared to the wild type enzyme. The mutant enzyme exclusively exhibits glycosynthase activity using alpha-D-glucosaminyl fluoride and D-glucosamine as substrates Photobacterium profundum
3.2.1.165 D139E compared with the wild type, the hydrolytic activity of the mutant is significantly suppressed (less than 0.1%), and the fluoride release activity also decreases to less than 3% Photobacterium profundum
3.2.1.165 D139S the mutation significantly decreases the hydrolytic activity compared to the wild type enzyme Photobacterium profundum
3.2.1.165 D143A the mutation significantly decreases the hydrolytic activity compared to the wild type enzyme Photobacterium profundum
3.2.1.165 E555A the mutation significantly decreases the hydrolytic activity compared to the wild type enzyme Photobacterium profundum

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.2.1.165 1.8
-
 chitobiose wild type enzyme, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 1.8
-
 chitobiose mutant enzyme D139E, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 3.5
-
 alpha-D-glucosaminyl fluoride mutant enzyme D139E, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 7.7
-
 chitobiose mutant enzyme D139A, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 15
-
 chitobiose mutant enzyme D139S, at pH 7.0 and 37°C Photobacterium profundum

Organism

EC Number Organism UniProt Comment Textmining
3.2.1.165 Photobacterium profundum Q6LUT2
-
-
3.2.1.165 Photobacterium profundum SS9 Q6LUT2
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.2.1.165 Ni-NTA column chromatography Photobacterium profundum

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.2.1.165 alpha-D-glucosaminyl fluoride + D-glucosamine
-
 Photobacterium profundum chitobiose + fluoride
-
 ?
3.2.1.165 alpha-D-glucosaminyl fluoride + D-glucosamine
-
 Photobacterium profundum SS9 chitobiose + fluoride
-
 ?
3.2.1.165 chitobiose + H2O
-
 Photobacterium profundum ?
-
 ?
3.2.1.165 chitobiose + H2O
-
 Photobacterium profundum SS9 ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.2.1.165 exo-beta-D-glucosaminidase
-
 Photobacterium profundum
3.2.1.165 GlcNase
-
 Photobacterium profundum

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.2.1.165 0.0047
-
 chitobiose mutant enzyme D139A, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.071
-
 chitobiose mutant enzyme D139E, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.073
-
 alpha-D-glucosaminyl fluoride mutant enzyme D139E, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.54
-
 chitobiose mutant enzyme D139S, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 87
-
 chitobiose wild type enzyme, at pH 7.0 and 37°C Photobacterium profundum

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.2.1.165 0.00062
-
 chitobiose mutant enzyme D139A, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.0019
-
 alpha-D-glucosaminyl fluoride mutant enzyme D139A, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.0031
-
 alpha-D-glucosaminyl fluoride mutant enzyme D139S, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.021
-
 alpha-D-glucosaminyl fluoride mutant enzyme D139E, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.036
-
 chitobiose mutant enzyme D139S, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.04
-
 chitobiose mutant enzyme D139E, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 0.73
-
 alpha-D-glucosaminyl fluoride wild type enzyme, at pH 7.0 and 37°C Photobacterium profundum
3.2.1.165 49
-
 chitobiose wild type enzyme, at pH 7.0 and 37°C Photobacterium profundum