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Information on EC 1.14.14.5 - alkanesulfonate monooxygenase and Organism(s) Escherichia coli and UniProt Accession P80645
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1.14.14.5 alkanesulfonate monooxygenaseIUBMB Comments
The enzyme from Escherichia coli catalyses the desulfonation of a wide range of aliphatic sulfonates (unsubstituted C1- to C14-sulfonates as well as substituted C2-sulfonates). Does not desulfonate taurine (2-aminoethanesulfonate) or aromatic sulfonates. Does not use FMN as a bound cofactor. Instead, it uses reduced FMN (i.e., FMNH2) as a substrate. FMNH2 is provided by SsuE, the associated FMN reductase (EC 1.5.1.38).
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Word Map
- 1.14.14.5
- flavin
- sulfur
-
desulfonation
-
two-component
- octanesulfonate
- sulfite
-
c4a-hydroperoxyflavin
-
organosulfonate
-
petroleum
-
oxygenolytic
-
tim-barrel
-
c4a-peroxyflavin
-
organosulfur
The taxonomic range for the selected organisms is: Escherichia coli
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea
Synonyms
alkanesulfonate monooxygenase, fmnh2-dependent alkanesulfonate monooxygenase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
alkanesulfonate alpha-hydroxylase
-
-
-
-
oxygenase, alkanesulfonate 1-mono-
-
-
-
-
SsuD
sulfate starvation-induced protein 6
-
-
-
-
SsuD
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
an alkanesulfonate + FMNH2 + O2 = an aldehyde + FMN + sulfite + H2O
absolutely dependent on oxygen
-
an alkanesulfonate + FMNH2 + O2 = an aldehyde + FMN + sulfite + H2O
catalysis by SsuD probably results in an unstable 1-hydroxysulfonate that spontaneously decomposes to the corresponding aldehyde and sulfite
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an alkanesulfonate + FMNH2 + O2 = an aldehyde + FMN + sulfite + H2O
kinetic reaction mechanism, charge transfer complex formation with flavin mononucleotide reductase in the alkanesulfonate monooxygenase system
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an alkanesulfonate + FMNH2 + O2 = an aldehyde + FMN + sulfite + H2O
mechanism of flavin reduction in the alkanesulfonate monooxygenase system, overview
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
desulfonation
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-
-
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SYSTEMATIC NAME
IUBMB Comments
alkanesulfonate,FMNH2:oxygen oxidoreductase
The enzyme from Escherichia coli catalyses the desulfonation of a wide range of aliphatic sulfonates (unsubstituted C1- to C14-sulfonates as well as substituted C2-sulfonates). Does not desulfonate taurine (2-aminoethanesulfonate) or aromatic sulfonates. Does not use FMN as a bound cofactor. Instead, it uses reduced FMN (i.e., FMNH2) as a substrate. FMNH2 is provided by SsuE, the associated FMN reductase (EC 1.5.1.38).
CAS REGISTRY NUMBER
COMMENTARY
54596-24-6
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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
octanesulfonate + FMNH2 + O2
octanal + FMN + sulfite + H2O
-
-
-
?
1,3-dioxo-2-isoindolineethanesulfonic acid + FMNH2 + O2
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)acetaldehyd + FMN + sulfite + H2O
-
-
-
-
?
2-(4-pyridyl)ethanesulfonic acid + FMNH2 + O2
pyridin-4-ylacetaldehyde
-
-
-
-
?
4-phenyl-1-butanesulfonic acid + FMNH2 + O2
4-phenylbutanol + FMN + sulfite + H2O
-
-
-
-
?
butanesulfonic acid + FMNH2 + O2
butanal + FMN + sulfite + H2O
-
-
-
-
?
decanesulfonic acid + FMNH2 + O2
decanal + FMN + sulfite + H2O
-
-
-
-
?
hexanesulfonic acid + FMNH2 + O2
hexanal + FMN + sulfite + H2O
-
-
-
-
?
N-phenyltaurine + FMNH2 + O2
anilinoacetaldehyde + FMN + sulfite + H2O
-
-
-
-
?
octanesulfonic acid + FMNH2 + O2
octanal + FMN + sulfite + H2O
-
-
-
-
?
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
-
-
-
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ir
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
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the enzyme is involved in scavenging sulfur from alkanesulfonates under sulfur starvation
-
-
?
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
-
the two-component alkanesulfonate monooxygenase system, with the flavin mononucleotide reductase, SsuE, being a part of it besides SsuD, utilizes reduced flavin as a substrate to catalyze a unique desulfonation reaction during times of sulfur starvation, protein-protein interactions are important in the mechanism of flavin transfer
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-
ir
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
-
the enzyme system is involved in scavenging sulfur from alkanesulfonates under sulfur starvation, overview
-
-
?
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
-
mechanism of flavin reduction in the alkanesulfonate monooxygenase system, the FMN reductase, SsuE, catalyzes the reduction of FMN by NADPH, and the reduced flavin is transferred to the monooxygenase, SsuD, overview
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-
?
octanesulfonate + FMNH2 + O2
octanal + FMN + sulfite + H2O
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SsuD shows a clear preference for FMNH2, reaction via C4a-(hydro)peroxyflavin intermediate
-
-
?
pentanesulfonic acid + FMNH2 + O2
pentaldehyde + FMN + sulfite + H2O
-
-
-
-
?
pentanesulfonic acid + FMNH2 + O2
pentaldehyde + FMN + sulfite + H2O
-
-
-
?
additional information
?
-
residues Arg226 donates a proton to the FMN-O? intermediate, triggering a conformational change that opens the enzyme to solvation and promotes product release, solvent and kinetic isotope studies
-
-
?
additional information
?
-
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further substrates: sulfoacetate, ethanesulfate, propanesulfonate, 2-hydroxyethanesulfonic acid, 3-aminopropanesulfate, no substrate: taurine
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-
?
additional information
?
-
-
no substrates are taurine, methanesulfonic acid, benzenesulfonic acid, L-cysteic acid, ethanedisulfonic acid, toluene-4-sulfonic acid, p-sulfobenzoic acid, benzenesulfonic acid, 4-hydroxybenzenesulfonic acid, SsuD is able to desulfonate C-2 to C-10 unsubstituted alkanesulfonates, substituted ethanesulfonic acids and HEPES, the catalytic efficiency increases with increasing chain length up to decanesulfonic acid
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-
?
additional information
?
-
-
mechanism of flavin reduction in the alkanesulfonate monooxygenase system, consisting of the alkanesulfonate monooxygenase and the flavin mononucleotide reductase, which catalyzes the reduction of FMN by NADPH, overview
-
-
?
additional information
?
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the enzyme interacts with the flavin mononucleotide reductase, SsuE, in a 1:1 monomeric association, mechanism of protein-protein interaction not leading to overall conformational changes in protein structure, overview
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-
?
additional information
?
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the two-component alkanesulfonate monooxygenase system from Escherichia coli includes an FMN reductase, SsuE, and an FMNH2-dependent alkanesulfonate monooxygenase, SsuD, involved in the acquisition of sulfur from alkanesulfonates during sulfur starvation, overview
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-
?
additional information
?
-
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Cys54 in SsuD may be either directly or indirectly involved in stabilizing the C4a-(hydro)peroxyflavin intermediate formed during catalysis through hydrogen bonding interactions
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-
?
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 two-component alkanesulfonate monooxygenase system from Escherichia coli includes an FMN reductase, SsuE, and an FMNH2-dependent alkanesulfonate monooxygenase, SsuD, involved in the acquisition of sulfur from alkanesulfonates during sulfur starvation, overview
-
-
?
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
-
the enzyme is involved in scavenging sulfur from alkanesulfonates under sulfur starvation
-
-
?
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
-
the two-component alkanesulfonate monooxygenase system, with the flavin mononucleotide reductase, SsuE, being a part of it besides SsuD, utilizes reduced flavin as a substrate to catalyze a unique desulfonation reaction during times of sulfur starvation, protein-protein interactions are important in the mechanism of flavin transfer
-
-
ir
an alkanesulfonate + FMNH2 + O2
an aldehyde + FMN + sulfite + H2O
-
the enzyme system is involved in scavenging sulfur from alkanesulfonates under sulfur starvation, overview
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0312
octanesulfonate
wild-type, pH 7.5, temperature not specified in the publication
0.0322
octanesulfonate
mutant E20A, pH 7.5, temperature not specified in the publication
0.0363
octanesulfonate
mutant D111A, pH 7.5, temperature not specified in the publication
additional information
additional information
-
kinetics of reaction of SsuD with FMNH2, rapid reaction and stopped flow kinetic analyses, dissociation constants for FMN and FMNH2, detailed overview
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additional information
additional information
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rapid reaction kinetic analysis of SsuE-catalyzed flavin reduction using NADPH
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.065
octanesulfonate
mutant E20A, pH 7.5, temperature not specified in the publication
0.675
octanesulfonate
mutant D111A, pH 7.5, temperature not specified in the publication
0.73
octanesulfonate
wild-type, pH 7.5, temperature not specified in the publication
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
18
octanesulfonate
mutant D111A, pH 7.5, temperature not specified in the publication
18
octanesulfonate
mutant E20A, pH 7.5, temperature not specified in the publication
23
octanesulfonate
wild-type, pH 7.5, temperature not specified in the publication
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
physiological function
optimal transfer of reduced flavin from NADPH-dependent FMN reductase SsuE to SsuD requires defined protein-protein interactions, but diffusion can occur under specified conditions. A SsuD variant containing substitutions of charged residues shows a 4fold decrease in coupled assays that include SsuE to provide reduced FMN, but there is no activity observed with an SsuD variant containing a deletion of the alpha-helix containing conserved charged amino acids
physiological function
salt bridges between Arg297 and Glu20 or Asp111 are not critical to the desulfonation mechanism. The predicted role of residue Arg297 is to favorably interact with the phosphate group of the reduced flavin. Arg226 functions as a protection group shielding FMNOO- from bulk solvent and is more pronounced when both FMNOO- and octanesulfonate are bound
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
molecular dynamics simulations for Ssud in substrate-free form, bound with FMNH2, bound with a C4a-peroxyflavin intermediate (FMNOO?), bound with octanesulfonate, cobound with FMNH2 and octanesulfonate, and cobound with FMNOO? and octanesulfonate
molecular dynamics simulations of wild-type SsuD and variant enzymes bound with different combinations of FMNH2, C4a-peroxyflavin intermediate FMNOO-, and octanesulfonate. Mobile loop conformations are open, closed, and semiclosed. The substrate-free SsuD system has a wide opening capable of providing full access for substrates to enter the active site. Upon binding FMNH2, SsuD adopts a closed conformation. Salt bridges, Asp111-Arg263 and Glu205-Arg271, are particularly important in maintaining the closed conformation. With both FMNH2 and octanesulfonate bound in SsuD, a second conformation is formed dependent upon a favorable pi-pi interaction between residues His124 and Phe261
X-ray characterization, tetramer 96 A x 90 A x 66 A, comprises two homodimers, monomer 60A x 50 A x 40 A, TIM-barrel protein
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D251A/D252A/E253A
mutation of conserved charged amino acids, 4fold decrease in kcat/Km value
DELTA251-261
deletion of alpha-helix containing conserved charged amino acids, complete loss of activity
R271A
mutation does not alter the catalytic activity, but in the presence of reduced flavin the variant is more susceptible to proteolytic digestion compared to wild-type
C54A
-
has little effect on FMN or FMNH2 binding, kcat/Km value decreases 6fold relative to wild-type
C54S
-
has little effect on FMN or FMNH2 binding, kcat/Km value increases 3fold relative to wild-type. Is able to generate the C4a-(hydro)peroxyflavin, but the rate of formation is increased 10fold relative to wild-type
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 15% glycerol, the activity increases slightly during the first 2 to 3 weeks of storage
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene ssuD, the alkanesulfonate monooxygenase system, expressed from the ssuEADCB operon, is comprised of a flavin reductase encoded by ssuE and monooxygenase encoded by ssuD, ssuD expressionin strain BL21(DE3)
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ligated into pET21a plasmid containing the ssuD gene, wild-type and mutants expressed in Escherichia coli BL21(DE3) super-competent cells
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Eichhorn, E.; van der Ploeg, J.R.; Leisinger, T.
Deletion analysis of the Escherichia coli taurine and alkanesulfonate transport systems
J. Bacteriol.
182
2687-2795
2000
Escherichia coli
Eichhorn, E.; van der Ploeg, J.R.; Leisinger, T.
Characterization of a two-component alkanesulfonate monooxygenase from Escherichia coli
J. Biol. Chem.
274
26639-26646
1999
Escherichia coli
Van der Ploeg, J.R.; Eichhorn, E.; Leisinger, T.
Sulfonate-sulfur metabolism and its regulation in Escherichia coli
Arch. Microbiol.
176
1-8
2001
Escherichia coli
Eichhorn, E.; Davey, C.A.; Sargent, D.F.; Leisinger, T.
Monooxygenase SsuD
J. Mol. Biol.
324
457-468
2002
Escherichia coli
Gao, B.; Ellis, H.R.
Altered mechanism of the alkanesulfonate FMN reductase with the monooxygenase enzyme
Biochem. Biophys. Res. Commun.
331
1137-1145
2005
Escherichia coli
Gao, B.; Ellis, H.R.
Mechanism of flavin reduction in the alkanesulfonate monooxygenase system
Biochim. Biophys. Acta
1774
359-367
2007
Escherichia coli
Abdurachim, K.; Ellis, H.R.
Detection of protein-protein interactions in the alkanesulfonate monooxygenase system from Escherichia coli
J. Bacteriol.
188
8153-8159
2006
Escherichia coli
Zhan, X.; Carpenter, R.A.; Ellis, H.R.
Catalytic importance of the substrate binding order for the FMNH2-dependent alkanesulfonate monooxygenase enzyme
Biochemistry
47
2221-2230
2008
Escherichia coli
Carpenter, R.A.; Zhan, X.; Ellis, H.R.
Catalytic role of a conserved cysteine residue in the desulfonation reaction by the alkanesulfonate monooxygenase enzyme
Biochim. Biophys. Acta
1804
97-105
2010
Escherichia coli
Robbins, J.; Ellis, H.
Steady-state kinetic isotope effects support a complex role of Arg226 in the proposed desulfonation mechanism of alkanesulfonate monooxygenase
Biochemistry
53
161-168
2014
Escherichia coli (P80645)
Armacost, K.; Musila, J.; Gathiaka, S.; Ellis, H.R.; Acevedo, O.
Exploring the catalytic mechanism of alkanesulfonate monooxygenase using molecular dynamics
Biochemistry
53
3308-3317
2014
Escherichia coli (P80645)
Dayal, P.V.; Singh, H.; Busenlehner, L.S.; Ellis, H.R.
Exposing the alkanesulfonate monooxygenase protein-protein interaction sites
Biochemistry
54
7531-7538
2015
Escherichia coli (P80645)
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