Information on EC 3.2.1.150 - oligoxyloglucan reducing-end-specific cellobiohydrolase

for references in articles please use BRENDA:EC3.2.1.150
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The expected taxonomic range for this enzyme is: Geotrichum

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EC NUMBER
COMMENTARY hide
3.2.1.150
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RECOMMENDED NAME
GeneOntology No.
oligoxyloglucan reducing-end-specific cellobiohydrolase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
Hydrolysis of cellobiose from the reducing end of xyloglucans consisting of a (1->4)-beta-linked glucan carrying alpha-D-xylosyl groups on O-6 of the glucose residues. To be a substrate, the first residue must be unsubstituted, the second residue may bear a xylosyl group, whether further glycosylated or not, and the third residue, which becomes the new terminus by the action of the enzyme, is preferably xylosylated, but this xylose residue must not be further substituted.
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of O-glycosyl bond
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
xyloglucan degradation III (cellobiohydrolase)
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SYSTEMATIC NAME
IUBMB Comments
oligoxyloglucan reducing-end cellobiohydrolase
The enzyme is found in the fungus Geotrichum sp. M128. The substrate is a hemicellulose found in plant cell walls.
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CAS REGISTRY NUMBER
COMMENTARY hide
37329-65-0
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753502-07-7
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
sp. M128
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Manually annotated by BRENDA team
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Uniprot
Manually annotated by BRENDA team
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SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
heptasaccharide XXXG
trisaccharide XG + tetrasaccharide XX
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?
oligoxyloglucan
?
show the reaction diagram
xyloglucan heptasaccharide
?
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?
xyloglucan tetradecasaccharide
?
show the reaction diagram
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enzyme substrate specificity to the tetradecasaccharide, XXXGXXXG, is analyzed. Wild-type OXG-RCBH generates XXXGXX and XG due to its strict specificity
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additional information
?
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no activity with beta-1,3/4-glucan and carboxymethyl cellulose (CMC)
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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
oligoxyloglucan
?
show the reaction diagram
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
15
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with xyloglucan heptasaccharide as substrate
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.5 - 5
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4.5
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assay at
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45
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assay at
50 - 60
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pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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PDB
SCOP
CATH
ORGANISM
UNIPROT
Geotrichum sp. (strain M128)
Geotrichum sp. (strain M128)
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MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
97000
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SDS-PAGE
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Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
by using the hanging-drop vapor-diffusion method. X-ray crystal structure of the OXG-RCBH-substrate complex is determined to a resolution of 2.4 A. OXG-RCBH consists of two seven bladed beta-propeller domains. There is a large cleft between the two domains, and a unique loop encloses one side of the active site cleft. Substrate bound to the cleft, and its reducing end is arranged near the loop region that is believed to impart OXG-RCBH with its activity
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at 2.2 A resolution
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hanging- and sitting drop vapor diffusion method with polyethylene glycol 3000 and polyethylene glycol 400 as precipitants
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 8
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stable at 45°C
646831
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
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10% loss of activity after 10 min at pH 4
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified by using a HiTrap DEAE FF column
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
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overexpressed in Escherichia coli BL21(DE3) as a His-tagged fusion protein
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D465N
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inactive OXG-RCBH is co-crystallized with a heptaxyloglucan substrate (XXXG) under conditions similar to the wild-type. There are no differences in the overall structure between the mutant and the wild-type
DELTAG375-H385
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a deltaloop mutant (deletion mutant of the loop region, Gly375–His385) is constructed. The mutant shows endo-activity with altered substrate recognition. More specifically, cleavage occurs randomly instead of at specific sites, most likely due to the misalignment of the substrate within the subsite. It is supposed that the loop imparts unique substrate specificity with exo-mode hydrolysis in OXG-RCBH
D35N
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inactive
D465N
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inactive
D490N
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inactive
E444Q
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inactive
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Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant protein is expressed as insoluble inclusion bodies. Purified inclusion bodies were solubilized in 8 M urea. Renaturing by dialysis results in enzymatically active enzyme
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LINKS TO OTHER DATABASES (specific for EC-Number 3.2.1.150)
ExplorEnz
ExPASy
KEGG
MetaCyc
SABIO-RK
NCBI: PubMed, Protein, Nucleotide, Structure, Genome, OMIM
IUBMB Enzyme Nomenclature
PROSITE Database of protein families and domains
InterPro (database of protein families, domains and functional sites)