Information on EC 3.2.1.145 - galactan 1,3-beta-galactosidase

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

EC NUMBER
COMMENTARY hide
3.2.1.145
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RECOMMENDED NAME
GeneOntology No.
galactan 1,3-beta-galactosidase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
hydrolysis of terminal, non-reducing beta-D-galactose residues in (1->3)-beta-D-galactopyranans
show the reaction diagram
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-
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-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of O-glycosyl bond
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SYSTEMATIC NAME
IUBMB Comments
galactan 3-beta-D-galactosidase
This enzyme removes not only free galactose, but also 6-glycosylated residues, e.g., (1->6)-beta-D-galactobiose, and galactose bearing oligosaccharide chains on O-6. Hence, it releases branches from [arabino-galacto-(1->6)]-(1->3)-beta-D-galactans.
CAS REGISTRY NUMBER
COMMENTARY hide
161515-48-6
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62213-17-6
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
subsp. longum
UniProt
Manually annotated by BRENDA team
subsp. longum
UniProt
Manually annotated by BRENDA team
gene Fo1,3 gal
UniProt
Manually annotated by BRENDA team
gene Fo1,3 gal
UniProt
Manually annotated by BRENDA team
strain NBRC 5367
-
-
Manually annotated by BRENDA team
recombinant enzyme
Swissprot
Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
NBCR14893
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
metabolism
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
alpha-L-arabinofuranosidase-treated root arabinogalactan proteins from radish + H2O
?
show the reaction diagram
relative activity: 9.3
-
-
?
beta-(1,3)-D-galactooligosaccharides + H2O
D-galactose + ?
show the reaction diagram
beta-(1,3)-galactobiose + H2O
D-galactose
show the reaction diagram
beta-(1-3)-D-galactan + H2O
beta-(1-3)-D-galactan + D-galactose
show the reaction diagram
beta-(1-3)-D-galactotriose + H2O
beta-(1-3)-D-galactobiose + D-galactose
show the reaction diagram
beta-(1-3)-galactobiose + H2O
D-galactose
show the reaction diagram
beta-(1-6)-galactopentaose + H2O
beta-(1-6)-D-galactotetraose + D-galactose
show the reaction diagram
50% of the activity with p-nitrophenyl-beta-galactoside
-
-
?
beta-(1-6)-galactotetraose + H2O
beta-(1-6)-D-galactotriose + D-galactose
show the reaction diagram
68% of the activity with p-nitrophenyl-beta-galactoside
-
-
?
beta-(1-6)-galactotriose + H2O
beta-(1-6)-D-galactobiose + D-galactose
show the reaction diagram
49% of the activity with p-nitrophenyl-beta-galactoside
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-
?
beta-1,3-beta-1,6-galactan + H2O
?
show the reaction diagram
relative activity: 3.1%
-
-
?
beta-1,3-D-galactobiose + H2O
2 D-galactose
show the reaction diagram
beta-1,3-D-galactobiose + H2O
D-galactose
show the reaction diagram
beta-1,3-D-galactopentaose + H2O
D-galactose + ?
show the reaction diagram
beta-1,3-D-galactotetraose + H2O
D-galactose + ?
show the reaction diagram
beta-1,3-D-galactotriose + H2O
D-galactose + ?
show the reaction diagram
-
-
-
?
beta-1,3-galactan + H2O
?
show the reaction diagram
beta-1,3-galactan + H2O
D-galactose
show the reaction diagram
beta-1,3-galactan + H2O
oligo-beta-(1->3)-D-galactose + D-galactose
show the reaction diagram
relative activity: 100%. Enzyme catalyzes hydrolysis only of beta-1,3-linked galactosyl oligosaccharides and polysaccharides. Enzyme does not hydrolyze beta-1,4-linked galactose-containing polysaccharides, such as pectic galactan and alpha-L-arabinofuranosidase-treated pectic galactan from lupine, and also does not hydrolyze laminarin, CM-curdlan, or beta-1,3-xylan, suggesting that the enzyme acts only on a beta-1,3-linked galactan chain
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-
?
beta-1,3-galactotriose + H2O
D-galactose + beta-(1,3)-galactobiose
show the reaction diagram
beta-D-Gal-[(1->3)-beta-D-Gal]n + H2O
beta-D-Gal-[(1->3)-beta-D-Gal]n-1 + D-galactose
show the reaction diagram
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only beta-(1, 3) oligomers act as substrates
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?
beta-galactosido-(1-3)-beta-galactosido-(1-6)-galactoside + H2O
beta-galactosido-(1-3)-beta-galactoside + D-galactose
show the reaction diagram
42% of the activity with p-nitrophenyl-beta-galactoside
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-
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beta-galactosido-(1-6)-beta-galactosido-(1-3)-galactoside + H2O
beta-galactosido-(1-6)-beta-galactoside + D-galactose
show the reaction diagram
40% of the activity with p-nitrophenyl-beta-galactoside
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dearabinosylated gum arabic + H2O
?
show the reaction diagram
dearabinosylated larch wood arabinogalactan + H2O
?
show the reaction diagram
best substrate
-
-
?
dyed de-arabinosylated gum arabic + H2O
?
show the reaction diagram
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preparation of de-arabinosylated arabinogalactan-(protein)s as substrates for determination of beta-galactanase activity, method development, covalent coupling of the RB5 dye to the substrate, overview
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?
gum arabic + H2O
?
show the reaction diagram
larch wood arabinogalactan + H2O
?
show the reaction diagram
larch wood arabinogalactan + H2O
D-galactose + beta-(1->6)-D-galactobiose + beta-(1->6)-D-galactotriose + O-alpha-L-arabinofuranosyl-(1->3)-O-beta-D-galactopyranosyl-(1->6)-O-beta-D-galactopyranosyl-(1->6)-D-galactopyranose + O-beta-D-galactopyranosyl-(1->6)-[O-alpha-L-arabinofuranosyl-(1->3)-]-O-beta-D-galactopyranosyl-(1->6)-D-galactopyranose + ?
show the reaction diagram
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the enzyme bypasses the branching points of beta-1,3-galactan backbones in larch wood arabinogalactan
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?
methyl-beta-1,3-D-galactopentaoside + H2O
methyl-beta-1,3-D-galactotetraoside + D-galactose
show the reaction diagram
methyl-beta-1,3-D-galactotetraoside + H2O
methyl-beta-1,3-D-galactotrioside + D-galactose
show the reaction diagram
p-nitrophenyl-beta-galactoside + H2O
p-nitrophenol + beta-galactose
show the reaction diagram
-
-
-
?
sugar beet debranched L-arabinan + H2O
?
show the reaction diagram
-
-
-
?
type II arabinogalactan + H2O
?
show the reaction diagram
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-
-
?
additional information
?
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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-(1-3)-D-galactan + H2O
beta-(1-3)-D-galactan + D-galactose
show the reaction diagram
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-
-
?
additional information
?
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Hg2+
1 mM, almost complete inhibition
iodoacetic acid
1 mM, about 80% inhibition
sodium dodecylsulfate
1 mM, about 80% inhibition
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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addition of galactose at a concentration of 2 mM to the reaction mixture does not affect the enzyme activity
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
31.6
beta-(1-3)-D-galactobiose
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0.77 - 19.9
beta-(1-3)-D-galactotriose
1.77
beta-(1-3)-galactobiose
pH 4.0, 37C
0.3
p-nitrophenyl-beta-galactoside
pH 4.0, 37C
additional information
additional information
kinetic parameters of enzyme BLLJ_1840 activity on different substrates, overview
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
32.5
beta-(1-3)-D-galactotriose
Raphanus sativus
Q6L619
pH 4.0, 37C
28.6
beta-(1-3)-galactobiose
Raphanus sativus
Q6L619
pH 4.0, 37C
123
p-nitrophenyl-beta-galactoside
Raphanus sativus
Q6L619
pH 4.0, 37C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0139
substrate beta-1,3-D-galactotetraose, pH 5.5, 40C
0.0157
substrate beta-1,3-D-galactopentaose, pH 5.5, 40C
0.0188
substrate beta-1,3-D-galactotriose, pH 5.5, 40C
0.0218
substrate beta-1,3-D-galactobiose, pH 5.5, 40C
0.113
substrate gum arabic, pH 5.5, 40C
0.88
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culture filtrate, using beta-(1-3)-D-galactan as substrate, in 20 mM Na phosphate buffer (pH 6.0) at 37C
1.01
substrate beta-1,3-galactan, pH 5.5, 40C
1.17
substrate larch wood arabinogalactan, pH 5.5, 40C
1.28
substrate dearabinosylated gum arabic, pH 5.5, 40C
1.7
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after 1.9fold purification, using beta-(1-3)-D-galactan as substrate, in 20 mM Na phosphate buffer (pH 6.0) at 37C
5
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substrate dyed de-arabinosylated gum arabic
16.2
substrate dearabinosylated larch wood arabinogalactan, pH 5.5, 40C
18.9
purified recombinant enzyme, pH 5.0, 30C, beta-1,3-galactan as substrate
21.3
beta-1,3-galactan as a substrate
40
purified native enzyme, pH 3.8, 48C; purified native enzyme, pH 5.0, 48C
47.6
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purified recombinant enzyme
64.9
pH 4.0, 37C
additional information
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3.7 units/mg, beta-1,3-galactan as a substrate; substrate beta-(1-3)-D-galactan: relative activity 100%; substrate beta-(1-3)-D-galactobiose: relative activity 7.6%; substrate beta-(1-3)-D-galactotriose: relative activity 57%; substrate methyl-beta-1,3-D-galactopentaoside: relative activity 100%; substrate methyl-beta-1,3-D-galactotetraoside: relative activity 99%
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2 - 6
almost inactive below and above
3 - 6.5
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activity range, pH profile, recombinant enzyme, overview
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10 - 55
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activity range, temperature profile, recombinant enzyme, overview
30 - 48
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retains full activity up to 48C, completely inactivated at 60C
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.45
about, sequence calculation; about, sequence calculation
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Bifidobacterium bifidum (strain S17)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
33654
x * 33654 + x * 44981, SDS-PAGE, MALDI-TOF and calculated
42000
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gel filtration
44981
x * 33654 + x * 44981, SDS-PAGE, MALDI-TOF and calculated
45000
x * 45000, about, SDS-PAGE, x * 48200, about, sequence calculation
45520
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x * 45520, mature enzyme, sequence calculation
47900
x * 47900, about, sequence calculation
48000
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x * 48000, SDS-PAGE
48200
x * 45000, about, SDS-PAGE, x * 48200, about, sequence calculation
51000
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native PAGE
58529
x * 64400, recombinant His-tagged enzyme, SDS-PAGE, x * 58529, sequence calculation
64400
x * 64400, recombinant His-tagged enzyme, SDS-PAGE, x * 58529, sequence calculation
106100
sequence calculation
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
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1 * 51000, SDS-PAGE
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
selenomethionine-labeled recombinant Pc1,3Gal43A, hanging-drop vapour-diffusion method, 25C, the reservoir solution contains 16% w/v PEG 10000, 95 mM ammonium sulfate, 95 mM Bis-Tris, pH 5.5, and 4.8% v/v glycerol, 3-10 days, X-ray diffraction structure determination and analysis at 1.8 A resolution, multiwavelength anomalous dispersion method, since molecular replacement is unsuccessful
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purified recombinant His-tagged wild-type and selenomethionine-labeled enzymes in complex with galactose, IPTG, galactan, and lactose, hanging drop vapor diffusion method, mixing of 0.001 ml of 50 mg/ml protein in a buffer containing 20 mM Tris-HCl, pH 8.0, and 100 mM NaCl, with 0.001 ml of reservoir solution containing 100 mM sodium acetate, pH 4.5, and 2.9-3.3 M sodium chloride,16C, 3 days, X-ray diffraction structure determination and analysis at 2.7-3.2 A resolution
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3 - 10
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more than 80% of the initial activity remains after 16 h of incubation at pH values from 3.0 to 10.0
713904
3.5 - 7.5
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after incubation for 24 h at 4C
171335
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 48
-
retains full activity up to 48C, completely inactivated at 60C
40
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the enzyme is stable at temperatures up to 40C
65
loss of 90% of activity
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, DEAE-Toyopearl 650-M column chromatography and Resource PHE column chromatography
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DEAE-cellulose, CM-Toyopearl, hydroylapatite, electrofocusing, purified from commercial driselase
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native enzyme 1.8fold from culture supernatant by ammonium sulfate fractionation, anion exchange chromatography, and gel filtration; native enzyme 1.8fold from culture supernatant by ammonium sulfate fractionation, anion exchange chromatography, and gel filtration
Ni-nitrilotriacetic acid agarose colum
Ni-NTA agarose column
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recombinant enzyme 0.7fold from Pichia pastoris cell culture medium by cation exchange chromatography and gel filtration
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recombinant enzyme expressed in Pichia pastoris
recombinant expression of His-tagged wild-type and selenomethionine-labeled enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration
recombinant selenmethionine-labeled Pc1,3Gal43A from Pichia pastoris by ultrafiltration, and hydrophobic interaction chromatography
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recombinant soluble extracellular enzyme 5.7fold from Pichia pastoris strain KM71H by anion exchange and hydrophobic interaction chromatography
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
BLLJ_1840, phylogenetic tree, expresion as soluble enzyme in Escherichia coli
DNA and amino acid sequence determination and analysis, expression of the gene encoding the enzyme fused to a yeast secretion signal sequence under control of the alcohol oxidase promoter in Pichia pastoris
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expressed in Escherichia coli
gene Fo1,3 gal, recombinant expression in Pichia pastoris strain KM71H, subcloning in Escherichia coli strain DH5alpha. The expression vector pPICZ?A contains the native Saccharomyces cerevisiae alpha-factor secretion signal, which allows Pichia pastoris to secrete heterologous proteins into the growth medium
Pc1,3Gal43A, expression in Pichia pastoris as selenmethionine-labeled protein
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recombinant expression of N-terminally His-tagged wild-type and selenomethionine-labeled enzymes in Escherichia coli strain BL21(DE3)
SGalase1, DNA and amino acid equence determination and analysis, sequence comparison, phylogenetic analysis and tree; SGalase2, DNA and amino acid equence determination and analysis, sequence comparison, phylogenetic analysis and tree
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
the enzyme is strongly induced by larch wood arabinogalactan
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E112A
site-directed mutagenesis, catalytically inactive mutant, the mutation eliminates the galactan hydrolysis activity of the enzyme while it does not disrupt its ligand binding ability
E112Q
site-directed mutagenesis, catalytically inactive mutant, the mutation eliminates the galactan hydrolysis activity of the enzyme while it does not disrupt its ligand binding ability
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis