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Information on EC 2.4.1.248 - cycloisomaltooligosaccharide glucanotransferase and Organism(s) Niallia circulans and UniProt Accession P94286

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EC Tree
     2 Transferases
         2.4 Glycosyltransferases
             2.4.1 Hexosyltransferases
                2.4.1.248 cycloisomaltooligosaccharide glucanotransferase
IUBMB Comments
Specific for (1->6)-alpha-D-glucans (dextrans) and, unlike cyclomaltodextrin glucanotransferase (EC 2.4.1.19), without activity towards (1->4)-alpha-D-glucans, such as amylose. It also has no activity on oligosaccharides, such as amylopectin and pullulan, containing (1->6)-alpha-D-glucosidic linkages at branch points. The enzyme from Bacillus circulans T-3040 has been shown to form cycloisomalto-oligosaccharides of three sizes (7, 8 and 9 glucose units). It will also catalyse the disproportionation of two isomalto-oligosaccharides molecules to yield a series of isomalto-oligosachharides and the addition of D-glucose to cycloisomalto-oligosaccharides with ring opening to form isomalto-oligosaccharides.
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Niallia circulans
UNIPROT: P94286
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Word Map
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The taxonomic range for the selected organisms is: Niallia circulans
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
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cyclizes part of a (1->6)-alpha-D-glucan chain by formation of a (1->6)-alpha-D-glucosidic bond
Synonyms
citase, cycloisomaltooligosaccharide glucanotransferase, citase-598k, citase-t3040, isocyclomaltooligosaccharide glucanotransferase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
isocyclomaltooligosaccharide glucanotransferase
-
-
SYSTEMATIC NAME
IUBMB Comments
(1-6)-alpha-D-glucan:(1-6)-alpha-D-glucan 6-alpha-D-[1-6alpha-D-glucano]-transferase (cyclizing)
Specific for (1->6)-alpha-D-glucans (dextrans) and, unlike cyclomaltodextrin glucanotransferase (EC 2.4.1.19), without activity towards (1->4)-alpha-D-glucans, such as amylose. It also has no activity on oligosaccharides, such as amylopectin and pullulan, containing (1->6)-alpha-D-glucosidic linkages at branch points. The enzyme from Bacillus circulans T-3040 has been shown to form cycloisomalto-oligosaccharides of three sizes (7, 8 and 9 glucose units). It will also catalyse the disproportionation of two isomalto-oligosaccharides molecules to yield a series of isomalto-oligosachharides and the addition of D-glucose to cycloisomalto-oligosaccharides with ring opening to form isomalto-oligosaccharides.
CAS REGISTRY NUMBER
COMMENTARY hide
156621-23-7
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
dextran
?
show the reaction diagram
-
-
-
?
soluble starch
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
-
-
?
soluble starch
cyclomaltooctaose
show the reaction diagram
-
main product
-
?
amylose
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
-
yield 27.2%
-
?
dextran
?
show the reaction diagram
-
-
synthesis of seven- to nine-glucose-membered cycloisomaltooligosaccharides
-
?
glycogen
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
-
yield 14.6%
-
?
maltoheptaose
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
-
yield 15.8%
-
?
maltohexaose
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
-
yield 12.7%
-
?
maltopentaose
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
-
yield 8.9%
-
?
maltotetraose
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
-
yield 4.3%
-
?
maltotriose
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
-
poor substrate
yield 0.9%
-
?
partially hydrolyzed starch
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
soluble starch
cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) + ?
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
additional information
?
-
CITase catalyzes the synthesis of cycloisomaltooligosaccharides with 7-17 glucose units from dextran
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Pb2+
-
1 mM, 130% of initial activity
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1-ethyl-3-[(3-dimethylamino)propyl]carbodiimide
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inactivation o both wild-type and mutants A452N and V744L, inactivation is reduced in presence of 10 mM Ca2+
EDTA
-
1 mM, 86% residual acitvity
Fe2+
-
1 mM, 94% residual acitvity
Fe3+
-
1 mM, 82% residual acitvity
Hg2+
-
1 mM, 55% residual acitvity
additional information
-
activity of free enzyme is not influenced by NaCl up to 5 M. The activity of enzyme immobilized on Chitopearl BCW-3505 is not influenced by NaCl up to 2 M
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
Michaelis-Menten kinetics for wild-type and deletion mutant enzymes, overview
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
14.2
recombinant CITase from Bacillus circulans strain G22-10, pH 5.5, 40°C
14.6
recombinant CITase from Bacillus subtilis strain 168 DELTAaprE DELTAnprE, pH 5.5, 40°C
6.1
-
pH 6.0, 45
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6
mutant MDELTA234
6.5
mutant MDELTA23DELTA
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
mutant MDELTA23DELTA
30
mutant MDELTA234
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5
-
isoelectric focusing
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
15 deletion mutant enzymes. M123DELTA (R4-deleted), MDELTA234 (R1-deleted), and MDELTA23DELTA (R1/R4-deleted) catalyze cycloisomaltooligosaccharide synthesis, but other mutants are inactive. M123DELTA, MDELTA234, and MDELTA23DELTA increase their Km values against dextran 40. The wild-type enzyme and M123DELTA produced cycloisomaltooligosaccharide-8 predominantly, but MDELTA234 and MDELTA23DELTA lose cycloisomaltooligosaccharide-8 production specificity. The kcat values of MDELTA234 and MDELTA23DELTA decrease, and these mutants show narrowed temperature and pH stability ranges
physiological function
CITase production is induced by dextran 40, isomaltose, isomaltotriose, and panose, and soluble starch but not by G67 or dextrin, which suggests that alpha-1,6 glucosidic linkages are required for CITase induction. Although CITase is induced by isomaltose, isomaltotriose, and panose, no cyclodextrans are produced in the culture. Cyclodextran-producing activity in the presence of soluble starch as the substrate is observed only in cultures containing dextran 40 or soluble starch. The production of CITase is significantly unaffected by glucose addition, but soluble starch-CITase activity almost completely disappears after glucose addition. A 135-kDa protein contributes to cyclodextran formation from starch in the presence of CITase
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION?
CTA1_NIACI
972
1
107431
Swiss-Prot
-
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
103000
recombinant CITAse, gel filtration
102071
x * 102071, calculated
106000
-
2 * 106000, SDS-PAGE
184000
-
gel filtration
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 102071, calculated
dimer
-
2 * 106000, SDS-PAGE
additional information
CITase contains an N-terminal conserved region of Ser1-Gly403, an insertion region R1 of Tyr404-Tyr492, two conserved regions R2 of Glu493-Ser596 and R3 Gly597-Met700, and a C-terminal variable region R4 of Lys701-Ser934
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
proteolytic modification
sequence encodes a signal peptide of 35 amino acids and a mature protein of 960 amino acids
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystal structure of the core fragment from Ser39 to Met738, devoid of its N-terminal signal peptide and C-terminal nonconserved regions. The structural model contains one catalytic (beta/alpha)8-barrel domain and three beta-domains. Domain N with an immunoglobulin-like beta-sandwich fold is attached to the N-terminus. Domain C with a Greek key beta-sandwich fold is located at the C-terminus, and a carbohydrate-binding module family 35 beta-jellyroll domain B is inserted between the 7thbeta-strand and the 7th alpha-helix of the catalytic domain A. The structures of the inactive catalytic nucleophile mutant enzyme complexed with isomaltohexaose, isomaltoheptaose, isomaltooctaose, and cycloisomaltooctaose reveal that the ligands bind in the catalytic cleft and the sugar-binding site of CBM35
crystals of CITase bearing an N-terminal His6 tag, resolution of 2.3 A, belong to space group P3121, containing a single molecule in the asymmetric unit.. Crystals of CITase bearing a C-terminal His6 tag, resolution of 1.9 A, belong to space group P212121, containing two molecules in the asymmetric unit
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
F268V/D469Y/A513V/Y515S
mutant produces three times as much megalo-cycloisomaltooligosaccharides (10-12 glucose units) and 1.5 times as much total cycloisomaltooligosaccharides (7-12 glucose units) as compared with the wild-type. The modified product size specificity is attributable to the construction of novel substrate-binding sites in the B-2 substrate-binding site and reactivity is improved by mutation on subsite -3 on the catalytic domain
A452N
-
3fold increase in reaction velocity, 9fold increase in Km value. Activation by Ca2+ and inactivation by Cu2+ are reduced
V744L
-
2fold increase in reaction velocity, 3fold increase in Km value. Activation by Ca2+ and inactivation by Cu2+ are reduced
additional information
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 10
wild-type enzyme and mutant M123DELTA, stable at
721720
5 - 9
stable at
721736
5.5 - 10
mutant MDELTA234, stable at
721720
6 - 8.5
mutant MDELTA23DELTA, stable at
721720
4.5 - 9
-
-
698519
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
35
mutant MDELTA23DELTA, stable up to
50
wild-type enzyme and mutant M123DELTA, stable up to
35
-
stable up to
40
-
in presence of 1 mM Ca2+, stable up to
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant extracellular CITase from protease-deficient Bacillus subtilis strain 168 by ammonium sulfate fractionation, anion exchange chromatography, and gel filtration and another step of anion exchange chromatography, recombinant extracellular CITase from Bacillus circulans G22-10 by ammonium sulfate fractionation, anion exchange chromatography, and gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
gene cit, expression in Escherichia coli
overexpression of CITase in Bacillus circulans strain G22-10, and, with the alpha-amylase promoter (PamyQ) and amyQ signal sequence of Bacillus amyloliquefaciens, in a protease-deficient Bacillus subtilis strain 168 as extracellular protein. The Bacillus subtilis host-vector system allows production of cycloisomaltooligosaccharides by direct fermentation of dextran along with high CITase production, which is not possible in Bacillus circulans G22-10 due to growth inhibition by dextran at high concentrations and limited production of CITase, stable expression up to 48 h
expression in Escherichia coli
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
CITase production is induced by dextran 40, isomaltose, isomaltotriose, and panose, and soluble starch but not by G67 or dextrin
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Kawamoto, H.; Oguma, T.; Sekine, H.; Kobayashi, M.
Utilization of NaCl to suppress the growth of contaminating microbes during the continuous production of cycloisomaltooligosaccharides by immobilized enzyme
Biochem. Eng. J.
12
161-164
2002
Niallia circulans
-
Manually annotated by BRENDA team
Watanabe, H.; Nishimoto, T.; Kubota, M.; Chaen, H.; Fukuda, S.
Cloning, sequencing, and expression of the genes encoding an isocyclomaltooligosaccharide glucanotransferase and an alpha -amylase from a Bacillus circulans strain
Biosci. Biotechnol. Biochem.
70
2690-2702
2006
Niallia circulans (A0P8W9), Niallia circulans
Manually annotated by BRENDA team
Watanabe, H.; Takakura-Yamamoto, R.; Kurose, M.; Yoshida, K.; Oku, K.; Sawatani, I.; Nishimoto, T.; Kubota, M.; Chaen, H.; Fukuda, S.
Production of isocyclomaltopentaose from starch using isocyclomaltooligosaccharide glucanotransferase
Biosci. Biotechnol. Biochem.
70
3013-3018
2006
Niallia circulans, Niallia circulans AM7
Manually annotated by BRENDA team
Kawakita, H.; Sugita, K.; Saito, K.; Tamada, M.; Sugo, T.; Kawamoto, H.
Production of cycloisomaltooligosaccharides from dextran using enzyme immobilized in multilayers onto porous membranes
Biotechnol. Prog.
18
465-469
2002
Niallia circulans, Niallia circulans T-3040
Manually annotated by BRENDA team
Kawamoto, H.; Oguma, T.; Sekine, H.; Kobayashi, M.
Immobilization of cycloisomaltooligosaccharide glucanotransferase for the production of cycloisomaltooligosaccharides from dextran
Enzyme Microb. Technol.
28
515-521
2001
Niallia circulans
Manually annotated by BRENDA team
Funane, K.; Nakai, S.; Terasawa, K.; Oguma, T.; Kawamoto, H.; Kitamura, Y.; Kobayashi, M.
Mutation of Bacillus cyclodextran glucanotransferase to increase its reaction velocity
J. Appl. Glycosci.
50
33-35
2003
Niallia circulans, Niallia circulans T-3040
-
Manually annotated by BRENDA team
Watanabe, H.; Nishimoto, T.; Chaen, H.; Fukuda, S.
A novel glucanotransferase that produces a cyclomaltopentaose cyclized by an alpha-1,6-linkage
J. Appl. Glycosci.
54
109-118
2007
Niallia circulans
-
Manually annotated by BRENDA team
Kawabata, Y.; Kimura, K.; Funane, K.
Extracellular production of cycloisomaltooligosaccharide glucanotransferase and cyclodextran by a protease-deficient Bacillus subtilis host-vector system
Appl. Microbiol. Biotechnol.
93
1877-1884
2012
Niallia circulans (P94286), Niallia circulans T-3040 (P94286)
Manually annotated by BRENDA team
Funane, K.; Kawabata, Y.; Suzuki, R.; Kim, Y.M.; Kang, H.K.; Suzuki, N.; Fujimoto, Z.; Kimura, A.; Kobayashi, M.
Deletion analysis of regions at the C-terminal part of cycloisomaltooligosaccharide glucanotransferase from Bacillus circulans T-3040
Biochim. Biophys. Acta
1814
428-434
2011
Niallia circulans (P94286), Niallia circulans T-3040 (P94286)
Manually annotated by BRENDA team
Suzuki, R.; Terasawa, K.; Kimura, K.; Fujimoto, Z.; Momma, M.; Kobayashi, M.; Kimura, A.; Funane, K.
Biochemical characterization of a novel cycloisomaltooligosaccharide glucanotransferase from Paenibacillus sp. 598K
Biochim. Biophys. Acta
1824
919-924
2012
Niallia circulans (P94286), Niallia circulans T-3040 (P94286), Paenibacillus sp. (G9MBW2)
Manually annotated by BRENDA team
Suzuki, N.; Kim, Y.M.; Momma, M.; Fujimoto, Z.; Kobayashi, M.; Kimura, A.; Funane, K.
Crystallization and preliminary X-ray crystallographic analysis of cycloisomaltooligosaccharide glucanotransferase from Bacillus circulans T-3040
Acta Crystallogr. Sect. F
69
946-949
2013
Niallia circulans (P94286), Niallia circulans T-3040 (P94286)
Manually annotated by BRENDA team
Funane, K.; Ichinose, H.; Araki, M.; Suzuki, R.; Kimura, K.; Fujimoto, Z.; Kobayashi, M.; Kimura, A.
Evidence for cycloisomaltooligosaccharide production from starch by Bacillus circulans T-3040
Appl. Microbiol. Biotechnol.
98
3947-3954
2014
Niallia circulans (P94286), Niallia circulans T-3040 (P94286)
Manually annotated by BRENDA team
Suzuki, R.; Suzuki, N.; Fujimoto, Z.; Momma, M.; Kimura, K.; Kitamura, S.; Kimura, A.; Funane, K.
Molecular engineering of cycloisomaltooligosaccharide glucanotransferase from Bacillus circulans T-3040: structural determinants for the reaction product size and reactivity
Biochem. J.
467
259-270
2015
Niallia circulans (P94286), Niallia circulans T-3040 (P94286)
Manually annotated by BRENDA team
Suzuki, N.; Fujimoto, Z.; Kim, Y.M.; Momma, M.; Kishine, N.; Suzuki, R.; Suzuki, S.; Kitamura, S.; Kobayashi, M.; Kimura, A.; Funane, K.
Structural elucidation of the cyclization mechanism of alpha-1,6-glucan by Bacillus circulans T-3040 cycloisomaltooligosaccharide glucanotransferase
J. Biol. Chem.
289
12040-12051
2014
Niallia circulans (P94286), Niallia circulans T-3040 (P94286)
Manually annotated by BRENDA team