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Information on EC 4.2.3.124 - 2-deoxy-scyllo-inosose synthase and Organism(s) Niallia circulans and UniProt Accession Q9S5E2
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4.2.3.124 2-deoxy-scyllo-inosose synthaseIUBMB Comments
Requires Co2+ . Involved in the biosynthetic pathways of several clinically important aminocyclitol antibiotics, including kanamycin, butirosin, neomycin and ribostamycin. Requires an NAD+ cofactor, which is transiently reduced during the reaction [1,4]. The enzyme from the bacterium Bacillus circulans forms a complex with the glutamine amidotransferase subunit of pyridoxal 5'-phosphate synthase (EC 4.3.3.6), which appears to stabilize the complex [6,7].
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Word Map
- 4.2.3.124
- aminoglycoside
- circulans
-
2-deoxystreptamine-containing
- 2-deoxystreptamine
-
carbocycle
- butirosin
-
aminocyclitols
- dehydroquinate
-
butirosin-producing
-
aglycon
- kanamycin
- neomycin
- tobramycin
-
actinomycete
- gentamicin
- nad+
- catechol
- tenebrarius
-
cyclization
- micromonospora
-
amacs
- aminoglycoside-aminocyclitols
-
benzenoid
- shikimate
- industry
The taxonomic range for the selected organisms is: Niallia circulans
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
2-deoxy-scyllo-inosose synthase, btrc2, doi synthase, alloh, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
DOI synthase
-
btrC
-
-
-
-
kanC
-
-
-
-
neoC
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
reaction mechanism, overview
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
reaction mechanism, overview
-
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
reaction mechanism, overview
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
catalyric reaction mechanism and substrate specificity, overview
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
DOIS catalyzes the multi-step and direct cyclization reaction of G-6-P consisting five crucial transformations, which include the initial NAD+-dependent dehydrogenation at C-4, the elimination of a phosphate from C-5 and C-6, the hydride comeback to C-4 to form a cyclic enol ether, and the finial aldol-type intramolecular condensation between C-6 and C-1
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
reaction and stereochemical mechanisms, overview
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
reaction mechanism of the multistep reaction, stereochemistry of the phosphate elimination reaction, proceeding through syn-elimination assisted by Glu 243, and the cyclization, an aldol condensation that proceeds via a boat conformation, overview
D-glucose 6-phosphate = 2-deoxy-L-scyllo-inosose + phosphate
the reaction mechanism includes oxidation of C-4, beta-elimination of phosphate, reduction of C-4, ring opening, and intramolecular aldol cyclization. Completely conserved E243 of BtrC is catalytically involved in the phosphate elimination step
PATHWAY SOURCE
PATHWAYS
BRENDA
-
-, -, -, -
SYSTEMATIC NAME
IUBMB Comments
D-glucose-6-phosphate phosphate-lyase (2-deoxy-scyllo-inosose-forming)
Requires Co2+ [2]. Involved in the biosynthetic pathways of several clinically important aminocyclitol antibiotics, including kanamycin, butirosin, neomycin and ribostamycin. Requires an NAD+ cofactor, which is transiently reduced during the reaction [1,4]. The enzyme from the bacterium Bacillus circulans forms a complex with the glutamine amidotransferase subunit of pyridoxal 5'-phosphate synthase (EC 4.3.3.6), which appears to stabilize the complex [6,7].
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
2-deoxy-D-glucose 6-phosphate
2,5-dideoxy-L-scyllo-inosose + phosphate
low activity
-
-
?
3-deoxy-D-glucose 6-phosphate
2,4-dideoxy-L-scyllo-inosose + phosphate
low activity
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
-
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
-
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
-
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
carbocycle formation
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
intramolecular carbocyclization
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
intramolecular carbocyclization, mechanism, overview
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
intramolecular carbocyclization, multistep process
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
multistep reaction
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
the enzyme converts ubiquitous D-glucose 6-phosphate into the specific carbocycle, 2-deoxy-scyllo-inosose with an aid of NAD+-NADH recycling
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
cyclization stereochemistry, overview
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
intramolecular carbocyclization, hydride abstraction and returning appeared to take place within the same glucose molecule
product identification by GC-MS analysis
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
pro-R stereospecific hydride transfer by NAD+-catalyzed 2-deoxy-scyllo-inosose synthase
-
-
?
additional information
?
-
the hydroxy group at C-4 is essential for enzyme activity, while the hydroxy groups at C-2 and C-3 are significant for efficial transformation of natural glucose 6-phosphate, but not essential for the reaction catalysis at the active site of enzyme, no activity with 4-deoxy-glucose 6-phosphate, substrate specificity, overview
-
-
?
additional information
?
-
specific hydrogen-bond accepting and donating interactions between the hydroxy groups at C-2 and C-3 of the substrate and the enzyme are responsible for the recognition
-
-
?
additional information
?
-
substrate specificity of 2-deoxy-scyllo-inosose synthase towards deoxyglucose 6-phosphates and proposed mechanism
-
-
?
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 hydroxy group at C-4 is essential for enzyme activity, while the hydroxy groups at C-2 and C-3 are significant for efficial transformation of natural glucose 6-phosphate, but not essential for the reaction catalysis at the active site of enzyme, no activity with 4-deoxy-glucose 6-phosphate, substrate specificity, overview
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
-
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
-
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
-
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
carbocycle formation
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
intramolecular carbocyclization
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
intramolecular carbocyclization, mechanism, overview
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
intramolecular carbocyclization, multistep process
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
multistep reaction
-
-
?
D-glucose 6-phosphate
2-deoxy-L-scyllo-inosose + phosphate
the enzyme converts ubiquitous D-glucose 6-phosphate into the specific carbocycle, 2-deoxy-scyllo-inosose with an aid of NAD+-NADH recycling
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
NAD+
essential for activity, not only used for the enzyme reaction but also stabilizing DOI synthase
NAD+
required, binding strutcure in presence of inhibitor carbaglucose 6-phosphate
NAD+
required, cycling via NADH in the multistep reaction process from D-glucose 6-phosphate to 2-deoxy-L-scyllo-inosose, overview
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Co2+
additional information
Co2+
essential for activity, not only used for the enzyme reaction but also stabilizing DOI synthase
additional information
the enzyme is not affected by Mg2+, Ca2+, Mn2+, and Fe2+
additional information
-
the enzyme is not affected by Mg2+, Ca2+, Mn2+, and Fe2+
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
carbaglucose-6-phosphate
a substrate analogue inhibitor, binding structure, overview. The inhibitor coordinates a cobalt ion in the active site
D-glucose 6-homophosphonate
competitive inhibition versus D-glucose 6-phosphate
D-glucose 6-phosphonate
competitive inhibition versus D-glucose 6-phosphate, hydride transfer from 6-phosphonate to NAD+
DL-carbaglucose 6-phosphate
a mechanism-based irreversible inhibitor, synthesis, overview. The alpha,beta-unsaturated intermediate traps a specific nucleophilic group in the active site through the Michael-type 1,4-addition. The covalently modified amino acid residue is Lys141
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.23
D-glucose 6-phosphate
pH 7.7, 35°C, recombinant wild-type enzyme, BtrC
0.23
D-glucose 6-phosphate
pH 7.7, 37°C, recombinant wild-type enzyme
0.42
D-glucose 6-phosphate
pH 7.7, 35°C, recombinant wild-type enzyme, BtrC + BtrC2
20
D-glucose 6-phosphate
above, pH 7.7, 37°C, recombinant mutant K141Q
additional information
additional information
steady state and pre-steady state kinetic analysis, overview
-
additional information
additional information
-
steady state and pre-steady state kinetic analysis, overview
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.007
D-glucose 6-phosphate
below, pH 7.7, 37°C, recombinant mutant K141Q
0.41
D-glucose 6-phosphate
pH 7.7, 37°C, recombinant wild-type enzyme
0.44
D-glucose 6-phosphate
pH 7.7, 35°C, recombinant wild-type enzyme, BtrC + BtrC2
1
D-glucose 6-phosphate
pH 7.7, 35°C, recombinant wild-type enzyme, BtrC
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.00035
D-glucose 6-phosphate
pH 7.7, 37°C, recombinant mutant K141Q
2.4
D-glucose 6-phosphate
pH 7.7, 37°C, recombinant wild-type enzyme
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.5 - 8.5
7.7
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
37
46
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
genes btrC and btrC2, the latter encoding the 20 kDa enzyme subunit
UniProt
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
additional information
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
malfunction
disruption of gene btrC2 reduces the growth rate and antibiotics production, the growth rate is restored by addition of NH4Cl, both by addition of yeast extract
metabolism
physiological function
additional information
evolution
2-deoxy-scyllo-inosose synthase, i.e. DOIS or BtrC, clearly belongs to the family of dehydroquinate synthase, DHQS
evolution
evolutionary relationship between dehydroquinate synthase and 2-deoxy-scyllo-inosose synthase, overview
metabolism
2-deoxy-scyllo-inosose synthase is the starter enzyme for 2-deoxystreptamine biosynthesis. 2-Deoxystreptamine is the aglycon of clinically important aminocyclitol antibiotics
metabolism
2-deoxy-scyllo-inosose synthase, DOIS, is a key enzyme in the biosynthesis of 2-deoxystreptamine-containing aminocyclitol antibiotics
metabolism
DOI synthase is the key starter enzyme of 2-deoxystreptamine biosynthesis
metabolism
key enzyme in the biosynthesis of 2-deoxystreptamine-containing aminocyclitol antibiotics
metabolism
key enzyme in the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics. Multistep cyclization of D-glucose-6-phosphate into the six-membered carbocycle 2-deoxy-L-scyllo-inosose is the first step of 2-deoxystreptamine biosynthesis
metabolism
key enzyme in the biosynthesis of 2-deoxytstreptamine. The function od BtrC2 in biosynthesis of butirosin is indirect. BtrC2 is involved in secondary as well as primary metabolism probably playing a role in stabilizing and regulating DOI synthase
metabolism
key enzyme in the biosynthesis of clinically important aminoglycoside antibiotics is 2-deoxy-scyllo-inosose synthase, which catalyzes carbocycle formation from D-glucose 6-phosphate to 2-deoxy-scyllo-inosose through a multistep reaction
metabolism
the enzyme is involved in the biosynthesis of another major group of classical aminocyclitol antibiotics which contain the 2-deoxystreptamine, DOS, moiety such as neomycin, kanamycin, tobramycin, gentamicin, sisomicin, butirosin, or ribostamycin
metabolism
-
the enzyme is involved in the biosynthesis of another major group of classical aminocyclitol antibiotics which contain the 2-deoxystreptamine, DOS, moiety such as neomycin, kanamycin, tobramycin, gentamicin, sisomicin, butirosin, or ribostamycin
physiological function
2-deoxy-scyllo-inosose synthase is involved in the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics and catalyzes the carbocyclic formation from D-glucose 6-phosphate into 2-deoxy-scyllo-inosose
physiological function
the DOI synthase reaction is a crucial starter step for the biosynthetic pathway of DOS-containing aminocyclitol antibiotics. The DOI synthase catalyzes the intramolecular carbocyclization of D-glucose 6-phosphate into the first non-aminogenous cyclitol 2-deoxy-L-scyllo-inosose, DOI
physiological function
the enzyme is crucial in the carbocyclization reaction as part of the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics. The DOI synthase catalyzes the intramolecular carbocyclization of D-glucose 6-phosphate into the first non-aminogenous cyclitol 2-deoxy-L-scyllo-inosose, DOI
physiological function
-
the enzyme is crucial in the carbocyclization reaction as part of the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics. The DOI synthase catalyzes the intramolecular carbocyclization of D-glucose 6-phosphate into the first non-aminogenous cyclitol 2-deoxy-L-scyllo-inosose, DOI
physiological function
the role of the 20 kDa non-catalytic protein BtrC2, associated with the DOI synthase, in butirosin biosynthesis is to stabilize the synthase by complexing as heterodimer with BtrC allowing the stable production of butirosin for long periods. BtrC2 is also involved in the biosynthesis of vitamin B6 in primary metabolism
physiological function
2-deoxy-scyllo-inosose (DOI) synthase, which uses glucose-6-phosphate as a substrate for DOI biosynthesis, is indispensably involved in the initial stage of the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics including butirosin, gentamicin, kanamycin, and tobramycin
physiological function
DOI synthase (DOIS) is responsible for the formation of DOI from D-glucose-6-phosphate in the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics such as neomycin and butirosin
additional information
active site structure is located between the N-terminal and C-terminal domains. Two subunits exist as a dimer in the asymmetric unit. The two active sites of the dimer are different. One contains a dephosphorylated compound derived from the inhibitor and the other includes the inhibitor without change. Comparison of active site between the native and complex BtrC, overview
additional information
-
active site structure is located between the N-terminal and C-terminal domains. Two subunits exist as a dimer in the asymmetric unit. The two active sites of the dimer are different. One contains a dephosphorylated compound derived from the inhibitor and the other includes the inhibitor without change. Comparison of active site between the native and complex BtrC, overview
additional information
the characteristic glutamate residue E243 of DOI synthase is a key determinant to distinguish the reaction mechanism between DOI synthase and dehydroquinate, DHQ, synthase as well as primary sequence, Substrate recognition models of DOI synthase and DHQ synthase, overview
additional information
-
the characteristic glutamate residue E243 of DOI synthase is a key determinant to distinguish the reaction mechanism between DOI synthase and dehydroquinate, DHQ, synthase as well as primary sequence, Substrate recognition models of DOI synthase and DHQ synthase, overview
additional information
while the reaction mechanisms are quite similar, 2-deoxy-scyllo-inosose synthase is distinct from 3-dehydroquinate synthase, EC 4.2.3.4, in the shikimate pathway, with respect to the quaternary structure, metal ion requirement, and the kinetic parameters
additional information
-
while the reaction mechanisms are quite similar, 2-deoxy-scyllo-inosose synthase is distinct from 3-dehydroquinate synthase, EC 4.2.3.4, in the shikimate pathway, with respect to the quaternary structure, metal ion requirement, and the kinetic parameters
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). MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
20000
1 * 40000, catalytic BtrC, + 1 * 20000, noncatalytic BtrC2, SDS-PAGE
23000
40000
1 * 40000, catalytic BtrC, + 1 * 20000, noncatalytic BtrC2, SDS-PAGE
40608
x * 40608, mass spectrometry, x * 40768, DL-carbaglucose 6-phosphate-enzyme complex, mass spectrometry
40746
1 * 42000, BtrC, + 1 * 23000, BtrC2, recombinant enzyme, SDS-PAGE, 1 * 40746, BtrC, sequence calculation
40768
x * 40608, mass spectrometry, x * 40768, DL-carbaglucose 6-phosphate-enzyme complex, mass spectrometry
42000
23000
1 * 42000, BtrC, + 1 * 23000, BtrC2, recombinant enzyme, SDS-PAGE, 1 * 40746, BtrC, sequence calculation
42000
1 * 42000, BtrC, + 1 * 23000, BtrC2, recombinant enzyme, SDS-PAGE, 1 * 40746, BtrC, sequence calculation
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
?
x * 40608, mass spectrometry, x * 40768, DL-carbaglucose 6-phosphate-enzyme complex, mass spectrometry
dimer
two subunits exist as a dimer in the asymmetric unit. The two active sites of the dimer are different. One contains a dephosphorylated compound derived from the inhibitor and the other includes the inhibitor without change
heterodimer
heterodimer
1 * 40000, catalytic BtrC, + 1 * 20000, noncatalytic BtrC2, SDS-PAGE
heterodimer
1 * 42000, BtrC, + 1 * 23000, BtrC2, recombinant enzyme, SDS-PAGE, 1 * 40746, BtrC, sequence calculation
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified free enzyme and enzyme in complex with substrate analogue inhibitor carbaglucose 6-phosphate, and with NAD+, and Co2+, hanging drop vapour diffusion method, mixing of 0.002 ml pf 4.2 mg/ml protein in 5 mM Tris-HCl, pH 7.7, and 0.2 mM CoCl2, with 0.002 ml of reservoir solution containing 40% w/v PEG 4000, 200 mM Li2SO4, and 100 mM Tris-HCl, pH 8.6, equilibration against 1 ml of reservoir solution, addition of 1 mM Co2+ and 1 mM inhibitor, 4°C, X-ray diffraction structure determination and analysis at 2.15-2.3 A resoltuion, heavy atom derivatization
purified recombinant enzyme, hanging drop vapour diffusion method, mixing of 0.002 ml pf 4.2 mg/ml protein in 5 mM Tris-HCl, pH 7.7, and 0.2 mM CoCl2, with 0.002 ml of reservoir solution containing 40% w/v PEG 4000, 200 mM Li2SO4, and 100 mM Tris-HCl, pH 8.6, equilibration against 1 ml of reservoir solution, 4°C, X-ray diffraction structure determination and analysis at 2.3 A resolution
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
E243Q
site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
K141Q
site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
additional information
additional information
complementation of a btrC2 disruption mutant by expression of wild-type btrC2 from plasmid pHB201 in Bacillus circulans
additional information
-
complementation of a btrC2 disruption mutant by expression of wild-type btrC2 from plasmid pHB201 in Bacillus circulans
additional information
disruption of gene btrC2 by insertion of a tetracycline resistance cassette
additional information
-
disruption of gene btrC2 by insertion of a tetracycline resistance cassette
additional information
introduction of DOIS gene btrC from Bacillus circulans and disruption of genes for phosphoglucose isomerase, D-glucose 6-phosphate dehydrogenase, and phosphoglucomutase, pgi, zwf and pgm, respectively, in Escherichia coli strain GI724. While GI724DELTAP/pGA-btrC and GI724DELTAPP/pGA-btrC possess functional Zwf and thus can catabolize glucose via the pentose phosphate pathway, glucose catabolism is completely inhibited in GI724DELTAPZ/pGA-btrC and GI724DELTAPZP/pGA-btrC due to the deletion of both pgi and zwf genes
additional information
-
introduction of DOIS gene btrC from Bacillus circulans and disruption of genes for phosphoglucose isomerase, D-glucose 6-phosphate dehydrogenase, and phosphoglucomutase, pgi, zwf and pgm, respectively, in Escherichia coli strain GI724. While GI724DELTAP/pGA-btrC and GI724DELTAPP/pGA-btrC possess functional Zwf and thus can catabolize glucose via the pentose phosphate pathway, glucose catabolism is completely inhibited in GI724DELTAPZ/pGA-btrC and GI724DELTAPZP/pGA-btrC due to the deletion of both pgi and zwf genes
additional information
construction of several metabolically engineered recombinant strains of Bacillus subtilis, and comparison of the DOI titer produced by the recombinants that express a heterologous DOI synthase. The expression of a natural btrC gene, encoding DOI synthase in butirosin-producing Bacillus circulans, in the heterologous host Bacillus subtilis strain 168-BSDOI-2 generates approximately 2.3 g/l 2-deoxy-scyllo-inosose (DOI), fed-batch fermentation, UPLC-ESI-MS/MS analysis of DOI production
additional information
transgenic production of 2-deoxy-scyllo-inosose (DOI) in cyanobacterium Synechococcus elongatus strain PCC 7942, production of 400 mg/l of DOI without using an external carbon source, method overview
additional information
-
transgenic production of 2-deoxy-scyllo-inosose (DOI) in cyanobacterium Synechococcus elongatus strain PCC 7942, production of 400 mg/l of DOI without using an external carbon source, method overview
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-75°C, purified enzyme, in Tris-HCl, pH 7.7, and 0.14 M NaCl, completely stable for 6 months
4°C- -75°C, purified recombinant enzyme, in 50 mM Tris-HCl buffer with 0.2 mM Co2+, loss of 60% activity within 10 days
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
native enzyme 148fold by ammonium sulfate fractionation, anion exchange and affinity chromatography, and gel filtration, followed by another different step of anion exchange chromatography
native enzyme by ammonium sulfate fractionation, dialysis and two different steps of anion exchange chromatography
recombinant BtrC from Escherichia coli by gel filtration and hydrophobic interaction chromatography
recombinant BtrC2 from Escherichia coli strain JM109 by ammonium sulfate fractionation, anion exchange chromatography, ultrafiltration, and gel filtration
recombinant enzyme by gel filtration, ultracentrifugation, and hydrophobic interaction chromatography
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
complementation of a btrC2 disruption mutant by expression of wild-type btrC2 from plasmid pHB201 in Bacillus circulans
gene brtC, functional recombinant expression in Bacillus subtilis strain BSDOI-2, and functional recombinant expression from codo-optimzed gene in Streptomyces tenebrarius
gene btrC, DNA and amino acid sequence determination and analysis, cloning in Escherichia coli strain JM 105, functional expression in Escherichia coli strain BL21(DE3)
gene btrC, functional recombinant expression in Synechococcus elongatus strain PCC 7942
gene DOIS, expression in Escherichia coli strain GI724 from plasmid pGA-btrC
genes btrC and btrC2, co-overexpression in Escherichia coli strain JM109
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
BtrC expression is highly reduced by supplementation with 0.01 mg/ml pyridoxal
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
industry
the enzyme converts the sustainable source glucose 6-phosphate to the carbocycle 2-deoxy-scylla-inosose, which easily aromatizes to yield catechol
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Nango, E.; Kumasaka, T.; Sato, T.; Tanaka, N.; Kakinuma, K.; Eguchi, T.
Crystallization and X-ray analysis of 2-deoxy-scyllo-inosose synthase, the key enzyme in the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics
Acta Crystallogr. Sect. F
61
709-711
2005
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2)
Huang, Z.; Kakinuma, K.; Eguchi, T.
Stereospecificity of hydride transfer in NAD+-catalyzed 2-deoxy-scyllo-inosose synthase, the key enzyme in the biosynthesis of 2-deoxystreptamine-containing aminocyclitol antibiotics
Bioorg. Chem.
33
82-89
2005
Niallia circulans (Q9S5E2)
Hirayama, T.; Kudo, F.; Huang, Z.; Eguchi, T.
Role of glutamate 243 in the active site of 2-deoxy-scyllo-inosose synthase from Bacillus circulans
Bioorg. Med. Chem.
15
418-423
2007
Niallia circulans (Q9S5E2), Niallia circulans
Iwase, N.; Kudo, F.; Yamauchi, N.; Kakinuma, K.
Substrate specificity of 2-deoxy-scyllo-inosose synthase, the starter enzyme for 2-deoxystreptamine biosynthesis, toward deoxyglucose-6-phosphates and proposed mechanism
Biosci. Biotechnol. Biochem.
62
2396-2407
1998
Niallia circulans (Q9S5E2), Niallia circulans SANK72073 (Q9S5E2)
Tamegai, H.; Nango, E.; Koike-Takeshita, A.; Kudo, F.; Kakinuma, K.
Significance of the 20-kDa subunit of heterodimeric 2-deoxy-scyllo-inosose synthase for the biosynthesis of butirosin antibiotics in Bacillus circulans
Biosci. Biotechnol. Biochem.
66
1538-1545
2002
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2)
Tamegai, H.; Sawada, H.; Nango, E.; Aoki, R.; Hirakawa, H.; Iino, T.; Eguchi, T.
Roles of a 20 kDa protein associated with a carbocycle-forming enzyme involved in aminoglycoside biosynthesis in primary and secondary metabolism
Biosci. Biotechnol. Biochem.
74
1215-1219
2010
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2)
Nango, E.; Kudo, F.; Eguchi, T.; Kakinuma, K.
Reaction stereochemistry of 2-deoxy-scyllo-inosose synthase, the key enzyme in the biosynthesis of 2-deoxystreptamine
Chem. Lett.
32
438-439
2003
Niallia circulans (Q9S5E2)
-
Kudo, F.; Yamauchi, N.; Suzuki, R.; Kakinuma, K.
Kinetic isotope effect and reaction mechanism of 2-deoxy-scyllo-inosose synthase derived from butirosin-producing Bacillus circulans
J. Antibiot.
50
424-428
1997
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2)
Kudo, F.; Tamegai, H.; Fujiwara, T.; Tagami, U.; Hirayama, K.; Kakinuma, K.
Molecular cloning of the gene for the key carbocycle-forming enzyme in the biosynthesis of 2-deoxystreptamine-containing aminocyclitol antibiotics and its comparison with dehydroquinate synthase
J. Antibiot.
52
559-571
1999
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2), Streptomyces fradiae, Streptomyces fradiae IFO 13147
Kudo, F.; Hosomi, Y.; Tamegai, H.; Kakinuma, K.
Purification and characterization of 2-deoxy-scyllo-inosose synthase derived from Bacillus circulans. A crucial carbocyclization enzyme in the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics
J. Antibiot.
52
81-88
1999
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2)
Kogure, T.; Wakisaka, N.; Takaku, H.; Takagi, M.
Efficient production of 2-deoxy-scyllo-inosose from D-glucose by metabolically engineered recombinant Escherichia coli
J. Biotechnol.
129
502-509
2007
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2)
Nango, E.; Eguchi, T.; Kakinuma, K.
Active site mapping of 2-deoxy-scyllo-inosose synthase, the key starter enzyme for the biosynthesis of 2-deoxystreptamine. Mechanism-based inhibition and identification of lysine-141 as the entrapped nucleophile
J. Org. Chem.
69
593-600
2004
Niallia circulans (Q9S5E2), Niallia circulans SANK72073 (Q9S5E2)
Nango, E.; Kumasaka, T.; Hirayama, T.; Tanaka, N.; Eguchi, T.
Structure of 2-deoxy-scyllo-inosose synthase, a key enzyme in the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics, in complex with a mechanism-based inhibitor and NAD+
Proteins
70
517-527
2008
Niallia circulans (Q9S5E2), Niallia circulans, Niallia circulans SANK72073 (Q9S5E2)
Watanabe, S.; Ozawa, H.; Kato, H.; Nimura-Matsune, K.; Hirayama, T.; Kudo, F.; Eguchi, T.; Kakinuma, K.; Yoshikawa, H.
Carbon-free production of 2-deoxy-scyllo-inosose (DOI) in cyanobacterium Synechococcus elongatus PCC 7942
Biosci. Biotechnol. Biochem.
82
161-165
2018
Niallia circulans (Q9S5E2), Niallia circulans
Lim, J.H.; Hwang, H.H.; Lee, N.J.; Lee, J.W.; Seo, E.G.; Son, H.B.; Kim, H.J.; Yoon, Y.J.; Park, J.W.
Enhanced biosynthesis of 2-deoxy-scyllo-inosose in metabolically engineered Bacillus subtilis recombinants
Front. Microbiol.
9
2333
2018
Niallia circulans (Q9S5E2), Streptoalloteichus tenebrarius (Q2MF16), Streptoalloteichus tenebrarius, Streptoalloteichus tenebrarius ATCC 17920 (Q2MF16)
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