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Information on EC 3.2.1.96 - mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase and Organism(s) Glutamicibacter protophormiae and UniProt Accession Q9ZB22
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3.2.1.96 mannosyl-glycoprotein endo-beta-N-acetylglucosaminidaseIUBMB Comments
A group of related enzymes.
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Word Map
- 3.2.1.96
- oligosaccharide
-
n-linked
- lysosomal
-
high-mannose
-
deglycosylation
- n-acetylglucosamine
- glycopeptides
- beta-glucuronidase
-
analbuminemic
- n-glycans
- alpha-mannosidase
- glycosidases
-
concanavalin
-
asparagine-linked
- tunicamycin
- mastitis
-
complex-type
- neuraminidase
-
pngase
-
exoglycosidases
-
transglycosylation
-
sialylated
- autolysins
- phytolacca
-
oxazoline
- swainsonine
- mucor
-
3.2.1.30
-
mannose-type
-
h-sensitive
- alpha-fucosidase
-
udder
-
biantennary
-
high-mannose-type
-
3hmannose
-
hybrid-type
-
intramammary
-
h-resistant
-
chitinolytic
-
lipid-linked
- 1-deoxymannojirimycin
- beta-n-acetyl-d-glucosaminidase
- glcnac2
- man8glcnac2
- synthesis
- nutrition
-
tetraantennary
- meningosepticum
- chitobiose
-
oligomannosidic
- analysis
-
holins
- medicine
- glc3man9glcnac2
- food industry
The taxonomic range for the selected organisms is: Glutamicibacter protophormiae
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
nagase, beta-n-acetylglucosaminidase, endo-beta-n-acetylglucosaminidase h, endo h, endo-beta-n-acetylglucosaminidase, murein hydrolase, endod, engase, endo-beta-n-acetylglucosaminidase f, endo-m, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
acetylglucosaminidase, endo-beta
-
-
-
-
di-N-acetylchitobiosyl beta-N-acetylglucosaminidase
-
-
-
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DI-N-acetylchitobiosyl beta-N-acetylglucosaminidase F1
-
-
-
-
DI-N-acetylchitobiosyl beta-N-acetylglucosaminidase F2
-
-
-
-
DI-N-acetylchitobiosyl beta-N-acetylglucosaminidase F3
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-
-
-
endo-beta-(1->4)-N-acetylglucosaminidase
-
-
-
-
endo-beta-acetylglucosaminidase
-
-
-
-
endo-beta-N-acetylglucosaminidase
endo-N-acetyl-beta-D-glucosaminidase
-
-
-
-
endo-N-acetyl-beta-glucosaminidase
-
-
-
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endoglycosidase F1
-
-
-
-
endoglycosidase F2
-
-
-
-
endoglycosidase F3
-
-
-
-
endoglycosidase S
-
-
-
-
mannosyl-glycoprotein 1,4-N-acetamidodeoxy-beta-D-glycohydrolase
-
-
-
-
Mannosyl-glycoprotein endo-beta-N-acetyl-glucosaminidase
-
-
-
-
Mannosyl-glycoprotein endo-beta-N-acetyl-glucosaminidase F1
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-
-
-
Mannosyl-glycoprotein endo-beta-N-acetyl-glucosaminidase F2
-
-
-
-
Mannosyl-glycoprotein endo-beta-N-acetyl-glucosaminidase F3
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-
-
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murein hydrolase
-
-
-
-
additional information
the enzyme belongs to the glycoside hydrolase family 85, GH85
endo-beta-N-acetylglucosaminidase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Endohydrolysis of the N,N'-diacetylchitobiosyl unit in high-mannose glycopeptides and glycoproteins containing the -[Man(GlcNAc)2]Asn- structure. One N-acetyl-D-glucosamine residue remains attached to the protein; the rest of the oligosaccharide is released intact
active site structure, catalytic mechanism in which residue E173 acts as the catalytic acid/base for reaction via an oxazoline intermediate, overview
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hydrolysis of O-glycosyl bond
-
-
-
-
PATHWAY SOURCE
PATHWAYS
SYSTEMATIC NAME
IUBMB Comments
glycopeptide-D-mannosyl-N4-(N-acetyl-D-glucosaminyl)2-asparagine 1,4-N-acetyl-beta-glucosaminohydrolase
A group of related enzymes.
CAS REGISTRY NUMBER
COMMENTARY
37278-88-9
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
GlcNAc-Asn + Manalpha(1-6)(Manalpha(1-3))Manbeta(1-4)GlcNAc
Manalpha(1-6)(Manalpha(1-3))Manbeta(1-4)GlcNAcbeta(1-4)GlcNAc-Asn
Manalpha(1-6)(Manalpha(1-3))Manbeta(1-4)GlcNAc-thiazoline is a transition state mimic and is resistant to Endo-A hydrolysis
-
-
?
(Man)6(GlcNAc)2-Asn + D-glucose
(Man)6GlcNAcGlc + ?
-
-
-
?
beta-Glcp-(1-4)-GlcpNAc-oxazoline
?
-
Endo-A shows only marginal activity for transglycosylation with the disaccharide oxazoline. When used in a relatively large quantity, Endo-A can promote the transglycosylation of the disaccharide oxazoline to a GlcpNAc-Asn acceptor (it catalyzes the transfer of alpha-Manp-(1-3)-beta-Glcp-(1-4)-GlcpNAc-oxazoline to the acceptor). Endo-A promotes polymerization of beta-Glcp-(1-4)-GlcpNAc-oxazoline
-
-
?
dansyl-Asn-(GlcNAc)2(Man)5 + H2O
dansyl-Asn-GlcNAc + ?
-
-
-
-
?
dansyl-Asn-(GlcNAc)2(Man)6 + H2O
dansyl-Asn-GlcNAc + (Man)6GlcNAc
-
-
-
-
?
O-(alpha-D-mannopyranosyl)-(1,6)-[(alpha-D-mannopyranosyl)-(1,3)]-beta-D-mannopyranosyl-(1,4)-(1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-oxazoline + benzyl alpha-D-mannopyranoside
2,3,4,6-tetra-O-acetyl-1-O-(2,2,2-trichloroethanimidoyl)-alpha-D-mannopyranose + ?
-
-
-
-
?
O-(beta-D-mannopyranosyl)-(1,4)-(1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-oxazoline + benzyl alpha-D-mannopyranoside
benzyl 2,4-di-O-benzoyl-alpha-D-mannopyranoside + ?
-
-
-
-
?
additional information
?
-
conserved essential catalytic residues E173, N171 and Y205 are within hydrogen bonding distance of the substrate. W216 and W244 regulate access to the active site during transglycosylation by serving as gate-keepers
-
-
?
additional information
?
-
-
conserved essential catalytic residues E173, N171 and Y205 are within hydrogen bonding distance of the substrate. W216 and W244 regulate access to the active site during transglycosylation by serving as gate-keepers
-
-
?
additional information
?
-
the enzyme is responsible for the hydrolysis of beta-1,4 linkage in the N,Ndiacetylchitobiose core of N-linked glycans. It also shows transglycosylation activity
-
-
?
additional information
?
-
-
the enzyme is responsible for the hydrolysis of beta-1,4 linkage in the N,Ndiacetylchitobiose core of N-linked glycans. It also shows transglycosylation activity
-
-
?
additional information
?
-
active site structure, catalytic mechanism in which residue E173 acts as the catalytic acid/base for reaction via an oxazoline intermediate, asparagine in the active centre is in a position likely to interact with the acetyl NH group. Three-dimensional structure of this important biocatalyst reveals that residues implicated in the enhancement of transglycosylation and synthetic capacity are proximal to the active centre, where they may act to favor binding of acceptor substrates
-
-
?
additional information
?
-
-
active site structure, catalytic mechanism in which residue E173 acts as the catalytic acid/base for reaction via an oxazoline intermediate, asparagine in the active centre is in a position likely to interact with the acetyl NH group. Three-dimensional structure of this important biocatalyst reveals that residues implicated in the enhancement of transglycosylation and synthetic capacity are proximal to the active centre, where they may act to favor binding of acceptor substrates
-
-
?
additional information
?
-
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the enzyme is probably devoted to exogeneous functions, such as degrading macromolecules for feeding purposes
-
-
?
additional information
?
-
-
enzyme has transglycosylation activity, high-mannose-type oligosaccharides are transferred to suitable glycosides as acceptor substrates, enzyme transfers high-mannose-type oligosaccharides more efficiently to beta-linked disaccharides than to alpha-linked disaccharides
-
-
?
additional information
?
-
-
the enzyme hydrolyzes the beta-1,4-glycosidic bond in the core N,N'-diacetylchitobiose moiety of N-glycoproteins to release the N-glycan. The enzyme also possesses transglycosylation activity and is able to transfer the released N-glycan to a GlcNAc-peptide acceptor to form a new glycopeptide
-
-
?
additional information
?
-
-
minimum structure of the donor substrate required for transglycosylation is the Man-beta-1,4-N-acetyl-Glc oxazoline moiety. Replacement of beta-D-mannose with beta-D-glucose, beta-D-galactose, or beta-D-N-acetylglucose monosaccharides results in loss of substrate activity for the disaccharide oxazoline
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
additional information
?
-
the enzyme is responsible for the hydrolysis of beta-1,4 linkage in the N,Ndiacetylchitobiose core of N-linked glycans. It also shows transglycosylation activity
-
-
?
additional information
?
-
-
the enzyme is responsible for the hydrolysis of beta-1,4 linkage in the N,Ndiacetylchitobiose core of N-linked glycans. It also shows transglycosylation activity
-
-
?
additional information
?
-
-
the enzyme is probably devoted to exogeneous functions, such as degrading macromolecules for feeding purposes
-
-
?
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
Manalpha(1-6)(Manalpha(1-3))Manbeta(1-4)GlcNAc-thiazoline
-
O-(2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl)-(1,6)-[(2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl)-(1,3)]-2,4-di-O-acetyl-beta-D-mannopyranosyl-(1,4)-(3,6-di-O-acetyl-1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-thiazoline
-
-
O-(alpha-D-mannopyranosyl)-(1,6)-[(alpha-D-mannopyranosyl)-(1,3)]-beta-D-mannopyranosyl-(1,4)-(1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-thiazoline
-
-
O-(beta-D-mannopyranosyl)-(1,4)-(1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-thiazoline
-
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.876
dansyl-Asn-(GlcNAc)2(Man)5
-
mutant E174A, presence of 2 M sodium azide, pH 6.0, 37°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0458
dansyl-Asn-(GlcNAc)2(Man)5
-
mutant E174A, presence of 2 M sodium azide, pH 6.0, 37°C
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.00022
O-(2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl)-(1,6)-[(2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl)-(1,3)]-2,4-di-O-acetyl-beta-D-mannopyranosyl-(1,4)-(3,6-di-O-acetyl-1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-thiazoline
Glutamicibacter protophormiae
-
pH 6.5, 30°C
0.013
O-(alpha-D-mannopyranosyl)-(1,6)-[(alpha-D-mannopyranosyl)-(1,3)]-beta-D-mannopyranosyl-(1,4)-(1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-thiazoline
Glutamicibacter protophormiae
-
pH 6.5, 30°C
0.038
O-(beta-D-mannopyranosyl)-(1,4)-(1,2-dideoxy-alpha-D-glucopyrano)-[2,1-d]-2-thiazoline
Glutamicibacter protophormiae
-
pH 6.5, 30°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.01
specific transglycosylation activity of mutant N171A
1.61
specific transglycosylation activity of mutant Y205F
1.72
specific transglycosylation activity of the wild-type
5.11
specific transglycosylation activity of mutant Y299F
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
-
Arthrobacter protophormiae strain RKJ100 employs the enzyme as an important component in osmotic shock response for resisting extreme concentrations of o-nitrobenzoate
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
the GH85 enzyme Endo-A displays a trimodular architecture in which a (betaalpha)8 catalytic domain occurs with two ancillary beta-sheet modules
additional information
-
the GH85 enzyme Endo-A displays a trimodular architecture in which a (betaalpha)8 catalytic domain occurs with two ancillary beta-sheet modules
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
Endo-A in the native form or in complex with Man3GlcNAc-thiazoline or GlcNAc-Asn, between 2-3.5 A resolution. Crystals of Endo-A belong to P1 space group with four molecules of the protein in the asymmetric unit. The carbohydrate moiety sits above the TIM-barrel in a cleft region surrounded by aromatic residues. N171 is hydrogen bonded to the thiazoline nitrogen, mimicking the ability of the asparagine to orient the acetamido group for a nucleophilic attack on the anomeric carbon
purifed recombinant wild-type enzyme and mutant E173Q in unlabeled or selenomethionine-labeled forms, 1:1 mixture of 42 mg/mL protein with a reservoir solution consisting of 0.35 M KSCN, 0.1 M 1,3-bis(tris(hydroxymethyl)methylamino)propane, pH 6.5, and 15% w/v PEG 3350, 30 days at 4 °C for the unlabeled, and 14 days at 18 °C for the SeM-labeled enzymes, X-ray diffraction and analysis at 1.8 A resolution, multiple-wavelength anomalous scattering methods, structure modelling, overview
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
E173Q
site-directed mutagenesis, the mutant shows increased transglycosylation activity compared to the wild-type enzyme, three-dimensional structure determination and analysis, overview
Y205F
exhibits reduced hydrolysis activity with an increase in transglycosylation yields
Y299F
3fold increase in the transglycosylation activity, while the hydrolysis activity remains unchanged
E173A
-
mutation in catalytically essential residue. Partial rescue of activity by addition of sodium azide or sodium formate, the produced beta-glycosyl azide retaines the anomeric configuration
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
on Ni-NTA resin and on ion-exchange chromatography column and by gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3), and of selenomethionine-labeled enzymes in strain B834(DE3)
into vector pET15b and transformed into Escherichia coli BL21(DE3) cells
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
analysis
-
essential reagent for the investigation of the structure and functions of glycoproteins
synthesis
additional information
-
polymerization by endo-beta-N-acetylglucosaminidase may find useful applications for the synthesis of novel artificial polysaccharides
synthesis
-
fluorescence-based assay for the transglycosylation activity of endo-beta-N-acetylglucosaminidases, highly sensitive, easy and quantitative method for screening endo-beta-N-acetylglucosaminidases with transglycosylation activity useful for glycoconjugate synthesis
synthesis
-
highly efficient chemoenzymatic synthesis of N-glycopeptides. The use of the synthetic oligosaccharide oxazolines as the donor substrates for the transglycosylation expands the substrate availability and results in substantial enhancement of the synthetic efficiency
synthesis
-
construction of natural or selectively modified glycopeptides by endoglycosidase-catalyzed transglycosylation
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Takegawa, K.; Yamabe, K.; Fujita, K.; Tabuchi, M.; Mita, M.; Izu, H.; Watanabe, A.; Asada, Y.; Sano, M.; Kondo, A.; Kato, I.; Iwahara, S.
Cloning, sequencing, and expression of Arthrobacter protophormiae endo-beta-N-acetylglucosaminidase in Escherichia coli
Arch. Biochem. Biophys.
338
22-28
1997
Glutamicibacter protophormiae
Karamanos, Y.; Bourgerie, S.; Barreaud, J.P.; Julien, R.
Are there biological functions for bacterial endo-N-acetyl-beta-D-glucosaminidases?
Res. Microbiol.
146
437-443
1995
Acinetobacter sp., Glutamicibacter protophormiae, Paenibacillus alvei, Elizabethkingia meningoseptica, Clostridium perfringens, Streptococcus pneumoniae, Flavobacterium sp., Myxococcus xanthus, Pseudomonas sp., Stigmatella aurantiaca, Streptomyces plicatus
Fujita, K.; Miyamura, T.; Sano, M.; Kato, I.; Takegawa, K.
Transfer of high-mannose-type oligosaccharides to disaccharides by endo-beta-N-acetylglucosaminidase from Arthrobacter protophormiae
J. Biosci. Bioeng.
93
614-617
2002
Glutamicibacter protophormiae
Hauser, S.; Song, H.; Li, H.; Wang, L.X.
A novel fluorescence-based assay for the transglycosylation activity of endo-beta-N-acetylglucosaminidases
Biochem. Biophys. Res. Commun.
328
580-585
2005
Glutamicibacter protophormiae, Mucor hiemalis
Li, B.; Zeng, Y.; Hauser, S.; Song, H.; Wang, L.
Highly efficient endoglycosidase-catalyzed synthesis of glycopeptides using oligosaccharide oxazolines as donor substrates
J. Am. Chem. Soc.
127
9692-9693
2005
Glutamicibacter protophormiae, Mucor hiemalis
Li, B.; Takegawa, K.; Suzuki, T.; Yamamoto, K.; Wang, L.X.
Synthesis and inhibitory activity of oligosaccharide thiazolines as a class of mechanism-based inhibitors for endo-beta-N-acetylglucosaminidases
Bioorg. Med. Chem.
15
4670-4675
2008
Glutamicibacter protophormiae, Flavobacterium sp., Homo sapiens, Mucor hiemalis
Zeng, Y.; Wang, J.; Li, B.; Hauser, S.; Li, H.; Wang, L.X.
Glycopeptide synthesis through endo-glycosidase-catalyzed oligosaccharide transfer of sugar oxazolines: probing substrate structural requirement
Chemistry
12
3355-3364
2006
Glutamicibacter protophormiae
Fujita, K.; Sato, R.; Toma, K.; Kitahara, K.; Suganuma, T.; Yamamoto, K.; Takegawa, K.
Identification of the catalytic acid base residue of Arthrobacter endo-beta-N-acetylglucosaminidase by chemical rescue of an inactive mutant
J. Biochem.
142
301-306
2007
Glutamicibacter protophormiae
Ochiai, H.; Huang, W.; Wang, L.X.
Endo-beta-N-acetylglucosaminidase-catalyzed polymerization of beta-Glcp-(1-->4)-GlcpNAc oxazoline: a revisit to enzymatic transglycosylation
Carbohydr. Res.
344
592-598
2009
Glutamicibacter protophormiae
Yin, J.; Li, L.; Shaw, N.; Li, Y.; Song, J.K.; Zhang, W.; Xia, C.; Zhang, R.; Joachimiak, A.; Zhang, H.C.; Wang, L.X.; Liu, Z.J.; Wang, P.
Structural basis and catalytic mechanism for the dual functional endo-beta-N-acetylglucosaminidase A
PLoS ONE
4
e4658
2009
Glutamicibacter protophormiae (Q9ZB22), Glutamicibacter protophormiae
Ling, Z.; Suits, M.D.; Bingham, R.J.; Bruce, N.C.; Davies, G.J.; Fairbanks, A.J.; Moir, J.W.; Taylor, E.J.
The X-ray crystal structure of an Arthrobacter protophormiae endo-beta-N-acetylglucosaminidase reveals a (beta/alpha)8 catalytic domain, two ancillary domains and active site residues key for transglycosylation activity
J. Mol. Biol.
389
1-9
2009
Glutamicibacter protophormiae (Q9ZB22), Glutamicibacter protophormiae
Pandey, J.; Khan, F.; Mahajan, V.; Pant, M.; Jain, R.K.; Pandey, G.
Evidence for vital role of endo-beta-N-acetylglucosaminidase in the resistance of Arthrobacter protophormiae RKJ100 towards elevated concentrations of o-nitrobenzoate
Extremophiles
18
491-500
2014
Glutamicibacter protophormiae, Glutamicibacter protophormiae RKJ100
EXTERNAL LINKS (specific for EC-Number 3.2.1.96)
Transporter Classification Database (TCDB): 1.C.105.2.7, 1.C.105.2.6
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