Information on EC 3.2.1.165 - exo-1,4-beta-D-glucosaminidase

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The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea

EC NUMBER
COMMENTARY hide
3.2.1.165
-
RECOMMENDED NAME
GeneOntology No.
exo-1,4-beta-D-glucosaminidase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
hydrolysis of chitosan or chitosan oligosaccharides to remove successive D-glucosamine residues from the non-reducing termini
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis
transglycosylation
-
only at high substrate concentrations
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Amino sugar and nucleotide sugar metabolism
-
-
Metabolic pathways
-
-
SYSTEMATIC NAME
IUBMB Comments
chitosan exo-(1->4)-beta-D-glucosaminidase
Chitosan is a partially or totally N-deacetylated chitin derivative that is found in the cell walls of some phytopathogenic fungi and comprises D-glucosamine residues with a variable content of GlcNAc residues [4]. Acts specifically on chitooligosaccharides and chitosan, having maximal activity on chitotetraose, chitopentaose and their corresponding alcohols [1]. The enzyme can degrade GlcN-GlcNAc but not GlcNAc-GlcNAc [3]. A member of the glycoside hydrolase family 2 (GH-2) [4].
CAS REGISTRY NUMBER
COMMENTARY hide
9012-33-3
c.f. EC 3.2.1.52
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain IAM2044
-
-
Manually annotated by BRENDA team
strain IAM2044
-
-
Manually annotated by BRENDA team
strain S-26
-
-
Manually annotated by BRENDA team
strain S-26
-
-
Manually annotated by BRENDA team
strain IAM2660
-
-
Manually annotated by BRENDA team
strain IAM2660
-
-
Manually annotated by BRENDA team
strain CJ22-326
UniProt
Manually annotated by BRENDA team
screening of natural sources and isolation and identification of Penicillium decumbens strain CFRNT15 with exo-beta-D-glucosaminidase activity
-
-
Manually annotated by BRENDA team
screening of natural sources and isolation and identification of Penicillium decumbens strain CFRNT15 with exo-beta-D-glucosaminidase activity
-
-
Manually annotated by BRENDA team
strain AF9-P-112
-
-
Manually annotated by BRENDA team
strain AF9-P-112
-
-
Manually annotated by BRENDA team
strain KY616
-
-
Manually annotated by BRENDA team
strain KY616
-
-
Manually annotated by BRENDA team
strain KOD1
SwissProt
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
4-methylumbelliferyl 2-amino-2-deoxy-beta-D-glucopyranoside + H2O
4-methylumbelliferone + 2-amino-2-deoxy-beta-D-glucopyranose
show the reaction diagram
-
-
-
-
?
4-methylumbelliferyl beta-D-glucoside + H2O
4-methylumbelliferol + D-glucose
show the reaction diagram
-
-
-
?
4-nitrophenyl beta-D-glucosaminide + H2O
4-nitrophenol + beta-D-glucosamine
show the reaction diagram
-
-
-
-
?
4-nitrophenyl beta-D-glucoside + H2O
4-nitrophenol + beta-D-glucose
show the reaction diagram
beta-1,4-D-glucosamine tetrasaccharide
D-glucosamine + chitotriose
show the reaction diagram
tetrasaccharide of chitosan efficiently depolymerizes chitosan to its constituent p-glucosamine monosaccharide in cooperation with endo-chitosanase
-
-
?
cellobiose + H2O
?
show the reaction diagram
cellopentaose + H2O
?
show the reaction diagram
cellotetraose + H2O
?
show the reaction diagram
-
-
-
-
?
cellotriose + H2O
?
show the reaction diagram
chitobiitol + H2O
?
show the reaction diagram
-
0.3% of the activity with chitopentaose
-
-
?
chitobiose + H2O
D-glucosamine
show the reaction diagram
chitobiose + H2O
glucosamine + glucosamine
show the reaction diagram
-
(GlcN)6 is sequentially cleaved into GlcN5,GlcN4, GlcN3, GlcN2, and GlcN, glucosamine is the final product
-
-
?
chitoheptaose + H2O
D-glucosamine + chitohexaose
show the reaction diagram
chitohexaitol + H2O
?
show the reaction diagram
-
94% of the activity with chitopentaose
-
-
?
chitohexaitol + H2O
D-glucosamine + ?
show the reaction diagram
chitohexaose + H2O
2-amino-2-deoxy-D-glucopyranose + chitopentaose
show the reaction diagram
-
GlcNn: polymerization degree of n=2-6, chitosan oligosaccharides, 0.96 nM exochiosanase, 10 mM ammonium acetate, pH 5.2, substrate concentrations: 6.25, 12.5, 25, 37.5 microM, hexa-N-acetylchitohexaose as control substrate at 25 microM
-
-
?
chitohexaose + H2O
chitopentaose + glucosamine
show the reaction diagram
-
(GlcN)6 is sequentially cleaved into GlcN5,GlcN4, GlcN3, GlcN2, and GlcN, glucosamine is the final product
-
-
?
chitohexaose + H2O
D-glucosamine
show the reaction diagram
chitohexaose + H2O
D-glucosamine + ?
show the reaction diagram
chitohexaose + H2O
D-glucosamine + chitopentaose
show the reaction diagram
chitopentaitol + H2O
?
show the reaction diagram
-
99% of the activity with chitopentaose
-
-
?
chitopentaitol + H2O
D-glucosamine + ?
show the reaction diagram
chitopentaose + H2O
?
show the reaction diagram
chitopentaose + H2O
chitotetraose + glucosamine
show the reaction diagram
-
(GlcN)6 is sequentially cleaved into GlcN5,GlcN4, GlcN3, GlcN2, and GlcN, glucosamine is the final product
-
-
?
chitopentaose + H2O
D-glucosamine
show the reaction diagram
chitopentaose + H2O
D-glucosamine + ?
show the reaction diagram
-
-
-
?
chitopentaose + H2O
D-glucosamine + chitotetraose
show the reaction diagram
chitosan + H2O
?
show the reaction diagram
chitosan + H2O
D-glucosamine
show the reaction diagram
chitosan + H2O
D-glucosamine + ?
show the reaction diagram
chitotetraitol + H2O
?
show the reaction diagram
-
as active as chitopentaose
-
-
?
chitotetraitol + H2O
D-glucosamine + ?
show the reaction diagram
chitotetraose + H2O
?
show the reaction diagram
chitotetraose + H2O
chitotriose + glucosamine
show the reaction diagram
-
(GlcN)6 is sequentially cleaved into GlcN5,GlcN4, GlcN3, GlcN2, and GlcN, glucosamine is the final product
-
-
?
chitotetraose + H2O
D-glucosamine
show the reaction diagram
chitotetraose + H2O
D-glucosamine + chitotriose
show the reaction diagram
-
products at an early stage of reaction
-
-
?
chitotriitol + H2O
?
show the reaction diagram
-
85% of the activity with chitopentaose
-
-
?
chitotriitol + H2O
D-glucosamine
show the reaction diagram
chitotriitol + H2O
D-glucosamine + ?
show the reaction diagram
-
85% of the activity with chitopentaitol
-
-
?
chitotriose + H2O
?
show the reaction diagram
-
-
-
?
chitotriose + H2O
chitobiose + D-glucosamine
show the reaction diagram
-
(GlcN)6 is sequentially cleaved into GlcN5,GlcN4, GlcN3, GlcN2, and GlcN, glucosamine is the final product
-
-
?
chitotriose + H2O
D-glucosamine
show the reaction diagram
chitotriose + H2O
D-glucosamine + chitobiose
show the reaction diagram
-
products at an early stage of reaction
-
-
?
deacetylated chitosan + H2O
?
show the reaction diagram
-
slight hydrolysis
-
-
?
GlcN(beta1-4)GlcN(beta1-4)GlcN + H2O
GlcN(beta1-4)GlcN + D-glucosamine
show the reaction diagram
-
-
-
-
?
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN + H2O
GlcN(beta1-4)GlcN(beta1-4)GlcN + D-glucosamine
show the reaction diagram
-
-
-
-
?
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN + H2O
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN + D-glucosamine
show the reaction diagram
-
-
-
-
?
GlcNbeta(1-4)GlcNAc + H2O
D-glucosamine + N-acetylglucosamine
show the reaction diagram
-
-
-
-
?
GlcNbeta(1-4)GlcNbeta(1-4)GlcNbeta(1-4)GlcNAc + H2O
GlcNbeta(1-4)GlcNbeta(1-4)GlcNAc + D-glucosamine
show the reaction diagram
glycol chitosan + H2O
?
show the reaction diagram
glycol chitosan + H2O
D-glucosamine + ?
show the reaction diagram
-
-
-
-
?
p-nitrophenyl beta-D-glucosaminide + H2O
p-nitrophenol + beta-D-glucosamine
show the reaction diagram
-
-
-
?
p-nitrophenyl-beta-D-glucosamine + H2O
D-glucosamine + p-nitrophenol
show the reaction diagram
-
-
-
?
p-nitrophenyl-beta-D-N-acetylglucosaminide + H2O
p-nitrophenol + beta-D-N-acetylglucosamine
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
beta-1,4-D-glucosamine tetrasaccharide
D-glucosamine + chitotriose
show the reaction diagram
Q56F26
tetrasaccharide of chitosan efficiently depolymerizes chitosan to its constituent p-glucosamine monosaccharide in cooperation with endo-chitosanase
-
-
?
chitosan + H2O
D-glucosamine
show the reaction diagram
additional information
?
-
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(1R,2R,3R,7S,7aR)-3-(aminomethyl)hexahydro-1H-pyrrolizine-1,2,7-triol
-
modest competitive inhibitor of CsxA
(1S,6S,7R,8R)-6-aminooctahydroindolizine-1,7,8-triol
-
potent competitive inhibitor of CsxA
2-Hydroxy-5-nitrobenzyl bromide
-
2.5 mM, complete inhibition
AlCl3
-
10 mM, 37% inhibition
CuSO4
-
10 mM, 55% inhibition
HgCl2
-
10 mM, complete inhibition
N-bromosuccinimide
-
10 mM, 34% inhibition
p-chloromercuribenzoic acid
-
2.5 mM, 86% inhibition
Pb(NO3)2
-
10 mM, 47% inhibition
ZnSO4
-
10 mM, 21% inhibition
additional information
-
neither castanospermine nor australine are capable of inhibiting the enzyme at a concentration of 1 mM
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.69
4-nitrophenyl beta-D-glucosaminide
-
pH 7.0, 37°C
4.5
4-nitrophenyl beta-D-glucoside
-
pH 7.0, 37°C
28.6
cellobiose
-
pH 7.0, 37°C
18.8
Cellotetraose
-
pH 7.0, 37°C
17.5
cellotriose
-
pH 7.0, 37°C
1.37 - 4.7
chitobiose
0.365
chitohexaose
pH 6.0, 80°C
0.0778
chitopentaose
pH 6.0, 80°C
0.15 - 0.295
chitotetraose
0.27 - 0.96
chitotriose
2.5
GlcN(beta1-4)GlcN(beta1-4)GlcN
-
pH 7.0, 37°C
1.2
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN
-
pH 7.0, 37°C
1.4
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN
-
pH 7.0, 37°C
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.087
4-nitrophenyl beta-D-glucosaminide
Photobacterium profundum
-
pH 7.0, 37°C
0.19
4-nitrophenyl beta-D-glucoside
Photobacterium profundum
-
pH 7.0, 37°C
68.8
cellobiose
Photobacterium profundum
-
pH 7.0, 37°C
54.2
Cellotetraose
Photobacterium profundum
-
pH 7.0, 37°C
47.7
cellotriose
Photobacterium profundum
-
pH 7.0, 37°C
152.9
chitobiose
Photobacterium profundum
-
pH 7.0, 37°C
27
chitohexaose
Amycolatopsis orientalis
-
2-amino-2-deoxy-D-glucopyranose split off, pH 5.2, 20°C
28.3
chitosan
Aspergillus fumigatus
-
-
77.4
GlcN(beta1-4)GlcN(beta1-4)GlcN
Photobacterium profundum
-
pH 7.0, 37°C
52
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN
Photobacterium profundum
-
pH 7.0, 37°C
97.6
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN
Photobacterium profundum
-
pH 7.0, 37°C
additional information
additional information
Amycolatopsis orientalis
Q56F26
kcat for high-molecular mass chitosan is 2832 per min
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.126
4-nitrophenyl beta-D-glucosaminide
Photobacterium profundum
-
pH 7.0, 37°C
41689
0.043
4-nitrophenyl beta-D-glucoside
Photobacterium profundum
-
pH 7.0, 37°C
712
2.4
cellobiose
Photobacterium profundum
-
pH 7.0, 37°C
82
2.5
Cellopentaose
Photobacterium profundum
-
pH 7.0, 37°C
767
2.9
Cellotetraose
Photobacterium profundum
-
pH 7.0, 37°C
590
2.7
cellotriose
Photobacterium profundum
-
pH 7.0, 37°C
516
32.6
chitobiose
Photobacterium profundum
-
pH 7.0, 37°C
1290
31.3
GlcN(beta1-4)GlcN(beta1-4)GlcN
Photobacterium profundum
-
pH 7.0, 37°C
28475
41.9
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN
Photobacterium profundum
-
pH 7.0, 37°C
28476
69.2
GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN(beta1-4)GlcN
Photobacterium profundum
-
pH 7.0, 37°C
41688
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.175
(1R,2R,3R,7S,7aR)-3-(aminomethyl)hexahydro-1H-pyrrolizine-1,2,7-triol
-
-
0.00061
(1S,6S,7R,8R)-6-aminooctahydroindolizine-1,7,8-triol
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.005
-
crude extract, using 4-nitrophenyl beta-D-glucoside as substrate, at pH 7.0, 37°C
0.046
-
purified enzyme, using 4-nitrophenyl beta-D-glucoside as substrate, at pH 7.0, 37°C
additional information
-
real-time MS as high sensitivity method to measure chitosanase activity quickly and with low substrate concentration, simultaneous detection of substrate and product
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.5 - 4.5
-
pH 3.5: about 60% of maximal activity, pH 4.5: about 50% of maximal activity
4 - 6
-
pH 4.0: about 55% of maximal activity, pH 6.0: about 60% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60 - 70
-
substrate: chitosan
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 60
-
30°C: about 70% of maximal activity, 60°C: about 45% of maximal activity
30 - 70
-
30°C: about 40% of maximal activity, 70°C: about 75% of maximal activity
37 - 100
37°C: about 20% of maximal activity, 100°C: about 20% of maximal activity
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
of cells grown in medium with chitosan
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45000
-
gel filtrration
86000
2 * 86000, SDS-PAGE
92000
-
gel filtration
93000
-
predicted molecular mass from atomic model, confirmed by SDS-PAGE of recombinant scxA gene product lacking carbohydrate-binding module
100000
105000
108000
-
x * 108000, SDS-PAGE
110000
-
1 * 110000, SDS-PAGE
110557
x * 110557, calculated from sequence
125000
-
gel filtration
135000
-
gel filtration
193000
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
2 * 86000, SDS-PAGE
monomer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
E541A mutant in complex with a natural β-1,4-D-glucosamine tetrasaccharide substrate and both E541A and D469A mutants in complex with a pNP-beta-D-glucosaminide synthetic substrate
-
hanging drop vapor diffusion method, using 14% (w/v) PEG 8000 and 0.2 M ammonium acetate in 0.1 M cacodylate buffer pH 7.0 for enzyme form Gls93-F1 or 20% (w/v) PEG 4000 and 0.2 M sodium isothiocyanate for enzyme form Gls93-F2
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2 - 10
-
stable
682639
3 - 7
-
stable
677420
3
-
1 h, 37°C, about 70% loss of activity
677651
3.5 - 7
-
37°C, 15 min, more than 50% of initial activity remains
681334
6 - 9
-
1 h, 37°C, stable
677651
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
-
pH 4.0, 1 h, stable
35
-
stable below
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C, pH 5.5, sodium acetate buffer, 4 months, stable
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, Q-Sepharose column chromatography, phenyl Sepharose column chromatography, and Sephacryl S-100 gel filtration
from culture supernatant
-
native enzyme by cation-exchange chromatography from chitosan-grown cells
-
Ni-NTA agarose column chromatography
-
Ni-NTA column chromatography and ultrafiltration
recombinant enzyme
SP-Sepharose ion exchange chromatography followed by hydroxyapatite column chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3) cells
-
expressed in Pichia pastoris
expression in Escherichia coli BL21
expression in Pichia pastoris
expression in Streptomyces lividans
-
Streptomyces lividans TK-24 with shuttle vector pFD666, M14 medium, pH 4.2
-
the His-tagged enzyme His-csx is expressed in Escherichia coli BL21(DE3) cells
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
the enzyme is induced by chitosan
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D469E
-
inactive mutant enzyme
E541A
-
catalytic nucleophile
E541D
-
inactive mutant enzyme
E541Q
-
inactive mutant enzyme
S468N/D469E
-
inactive mutant enzyme
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis
Show AA Sequence (238 entries)
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