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Literature summary extracted from

  • Tsuji, H.; Nishimura, S.; Inui, T.; Kado, Y.; Ishikawa, K.; Nakamura, T.; Uegaki, K.
    Kinetic and crystallographic analyses of the catalytic domain of chitinase from Pyrococcus furiosus - the role of conserved residues in the active site (2010), FEBS J., 277, 2683-2695.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.2.1.14 overexpression of wild-type enzyme and mutant enzymes D522N, D522A, D524N, D524A, E526Q and E526A in Escherichia coli Pyrococcus furiosus

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
3.2.1.14 hanging drop vapor diffusion method, crystallographic analyses of the catalytic domain of chitinase. Crystals of these mutant enzymes E526A and D524A in complex with chito-oligosaccharide substrate by means of cocrystallization and soaking methods, respectively, and determination of their tertiary structures. X-ray diffraction data are collected, resolutions of 1.80 A and 1.76 A Pyrococcus furiosus

Protein Variants

EC Number Protein Variants Comment Organism
3.2.1.14 D522A mutation causes a 7.3fold decrease in kcat/Km for 4-nitrophenyl chitobioside Pyrococcus furiosus
3.2.1.14 D522N mutation causes a 3.4fold decrease in kcat/Km for 4-nitrophenyl chitobioside Pyrococcus furiosus
3.2.1.14 D524A mutation causes a 58.4 fold decrease in kcat/Km for 4-nitrophenyl chitobioside Pyrococcus furiosus
3.2.1.14 D524N mutation causes a 3.6fold decrease in kcat/Km for 4-nitrophenyl chitobioside Pyrococcus furiosus
3.2.1.14 E526A no activity detected with 4-nitrophenyl chitobioside Pyrococcus furiosus
3.2.1.14 E526Q mutation causes a 38.4fold decrease in kcat/Km for 4-nitrophenyl chitobioside Pyrococcus furiosus
3.2.1.14 W664A mutation causes a 8588fold decrease in kcat/Km for 4-nitrophenyl chitobioside Pyrococcus furiosus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.2.1.14 0.09
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D524A Pyrococcus furiosus
3.2.1.14 0.12
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme E526Q Pyrococcus furiosus
3.2.1.14 0.46
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, wild-type enzyme Pyrococcus furiosus
3.2.1.14 0.54
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D522N Pyrococcus furiosus
3.2.1.14 0.61
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D524N Pyrococcus furiosus
3.2.1.14 0.74
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D522A Pyrococcus furiosus
3.2.1.14 12.7
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme W664A Pyrococcus furiosus

Organism

EC Number Organism UniProt Comment Textmining
3.2.1.14 Pyrococcus furiosus Q8U1H5
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.2.1.14
-
 Pyrococcus furiosus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.2.1.14 4-nitrophenyl beta-N,N'-diacetylchitobioside + H2O
-
 Pyrococcus furiosus 4-nitrophenol + N,N'-diacetylchitobiose
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.2.1.14 PF1233 locus name Pyrococcus furiosus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.2.1.14 50
-
 assay at Pyrococcus furiosus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.2.1.14 0.022
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D524A Pyrococcus furiosus
3.2.1.14 0.022
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme W664A Pyrococcus furiosus
3.2.1.14 0.045
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme E526Q Pyrococcus furiosus
3.2.1.14 1.49
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D522A Pyrococcus furiosus
3.2.1.14 2.36
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D522N Pyrococcus furiosus
3.2.1.14 2.47
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, mutant enzyme D524N Pyrococcus furiosus
3.2.1.14 6.7
-
 4-nitrophenyl beta-N,N'-diacetylchitobioside pH 4.8, 50°C, wild-type enzyme Pyrococcus furiosus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.2.1.14 4.8
-
 assay at Pyrococcus furiosus

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.2.1.14 0.0017
-
 4-nitrophenyl chitobioside pH 4.8, 50°C, mutant enzyme W664A Pyrococcus furiosus
3.2.1.14 0.25
-
 4-nitrophenyl chitobioside pH 4.8, 50°C, mutant enzyme D524A Pyrococcus furiosus
3.2.1.14 0.38
-
 4-nitrophenyl chitobioside pH 4.8, 50°C, mutant enzyme E526Q Pyrococcus furiosus
3.2.1.14 2
-
 4-nitrophenyl chitobioside pH 4.8, 50°C, mutant enzyme D522A Pyrococcus furiosus
3.2.1.14 4.1
-
 4-nitrophenyl chitobioside pH 4.8, 50°C, mutant enzyme D524N Pyrococcus furiosus
3.2.1.14 4.3
-
 4-nitrophenyl chitobioside pH 4.8, 50°C, mutant enzyme D522N Pyrococcus furiosus
3.2.1.14 14.6
-
 4-nitrophenyl chitobioside pH 4.8, 50°C, wild-type enzyme Pyrococcus furiosus