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Information on EC 1.11.1.23 - (S)-2-hydroxypropylphosphonic acid epoxidase and Organism(s) Streptomyces wedmorensis and UniProt Accession Q56185
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1.11.1.23 (S)-2-hydroxypropylphosphonic acid epoxidaseIUBMB Comments
This is the last enzyme in the biosynthetic pathway of fosfomycin, a broad-spectrum antibiotic produced by certain Streptomyces species. Contains non heme iron that forms a iron(IV)-oxo (ferryl) complex with hydrogen peroxide, which functions as a proton abstractor from the substrate .
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The taxonomic range for the selected organisms is: Streptomyces wedmorensis
The expected taxonomic range for this enzyme is: Bacteria, Archaea, Eukaryota
The expected taxonomic range for this enzyme is: Bacteria, Archaea, Eukaryota
Reaction Schemes
Synonyms
hpp epoxidase, (s)-2-hydroxypropylphosphonic acid epoxidase, ps-hppe, 2-hydroxypropylphosphonic acid epoxidase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
HppE
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(S)-2-hydroxypropylphosphonate + H2O2 = (1R,2S)-1,2-epoxypropylphosphonate + 2 H2O
a straightforward epoxidase mechanism is proposed that depends on the Lewis acid properties of divalent cations and the redox properties of FMN
(S)-2-hydroxypropylphosphonate + H2O2 = (1R,2S)-1,2-epoxypropylphosphonate + 2 H2O
an iron-redox mechanism is proposed in which the active site bound Fe2+ serves as a Lewis acid to activate the 2-OH group of (S)-2-hydroxypropylphosphonic acid, and the epoxide ring is formed by the attack of the 2-OH group at C-1 coupled with the transfer of the C-1 hydrogen as a hydride ion to the bound FMN
(S)-2-hydroxypropylphosphonate + H2O2 = (1R,2S)-1,2-epoxypropylphosphonate + 2 H2O
Tyr105 is a key residue, which plays a role in the activation of dioxygen required for the enzymatic activity of HppE
PATHWAY SOURCE
PATHWAYS
-
-
SYSTEMATIC NAME
IUBMB Comments
(S)-2-hydroxypropylphosphonate:hydrogen peroxide epoxidase
This is the last enzyme in the biosynthetic pathway of fosfomycin, a broad-spectrum antibiotic produced by certain Streptomyces species. Contains non heme iron that forms a iron(IV)-oxo (ferryl) complex with hydrogen peroxide, which functions as a proton abstractor from the substrate [7].
CAS REGISTRY NUMBER
COMMENTARY
261769-38-4
-
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
(3,3,3-trifluoro-1-hydroxypropyl)phosphonic acid + H2O2
?
-
-
-
?
(R)-2-hydroxypropylphosphonate + H2O2
2-oxopropylphosphonic acid + H2O
-
-
-
?
(R)-2-hydroxypropylphosphonic acid + H2O2
2-oxo-propylphosphonic acid + H2O
-
-
-
?
(S)-2-hydroxypropylphosphonate + H2O2
(1R,2S)-1,2-epoxypropylphosphonate + H2O
-
-
-
?
(S)-2-hydroxypropylphosphonate + H2O2
cis-fosfomycin + trans-fosfomycin + 2 H2O
-
-
-
?
(S)-2-hydroxypropylphosphonic acid + 2 NADH + O2
cis-(1R,2S)-epoxypropylphosphonic acid + 2 H2O + 2 NAD+
[(1R)-3,3,3-trifluoro-1-hydroxypropyl]phosphonate + H2O2
?
-
-
-
?
[(2R)-2-cyclopropyl-2-hydroxyethyl]phosphonate + H2O2
?
-
-
-
?
[(2S)-2-cyclopropyl-2-hydroxyethyl]phosphonate + H2O2
?
-
-
-
?
[(2S)-2-hydroxy-2-(2-methylidenecyclopropyl)ethyl]phosphonate + H2O2
?
-
-
-
?
[3-(2-methylidenecyclopropyl)oxiran-2-yl]phosphonate + H2O2
?
-
-
-
?
(R)-2-hydroxypropylphosphonate + H2O2
2-oxo-propylphosphonic acid + H2O
-
-
-
-
?
(R)-2-hydroxypropylphosphonate + NADH + H+ + O2
2-oxopropylphosphonic acid + NAD+ + H2O
-
-
-
-
?
(S)-2-hydroxypropylphosphonate + H2O2
(1R,2S)-1,2-epoxypropylphosphonate + H2O
-
-
-
-
?
(S)-2-hydroxypropylphosphonate + NADH + H+ + O2
(1R,2S)-epoxypropylphosphonate + NAD+ + H2O
-
-
i.e. fosfomycin
-
?
[(2R)-2-hydroxy-2-[(1R)-2-methylidenecyclopropyl]ethyl]phosphonate + H2O2
?
-
-
-
-
?
[(2R)-2-hydroxy-2-[(1S)-2-methylidenecyclopropyl]ethyl]phosphonate + H2O2
?
-
-
-
-
?
[(2R,3S)-3-[(1R)-2-methylidenecyclopropyl]oxiran-2-yl]phosphonate + H2O2
?
-
-
-
-
?
[(2R,3S)-3-[(1S)-2-methylidenecyclopropyl]oxiran-2-yl]phosphonate + H2O2
?
-
-
-
-
?
[(2S)-2-hydroxy-2-[(1R)-2-methylidenecyclopropyl]ethyl]phosphonic acid + H2O2
?
-
-
-
-
?
[2-[(1S)-2-methylidenecyclopropyl]-2-oxoethyl]phosphonate + H2O2
?
-
-
-
-
?
additional information
?
-
the enzyme also forms 2-oxopropylphosphonic acid from (R)-2-hydroxypropylphosphonic acid. HPP epoxidase is not selective with regard to substrate recognition, but can stereospecifically convert each enantiomer into a unique product with similar efficiency
-
-
?
(S)-2-hydroxypropylphosphonic acid + 2 NADH + O2
cis-(1R,2S)-epoxypropylphosphonic acid + 2 H2O + 2 NAD+
-
i.e. fosfomycin
-
?
(S)-2-hydroxypropylphosphonic acid + 2 NADH + O2
cis-(1R,2S)-epoxypropylphosphonic acid + 2 H2O + 2 NAD+
-
-
i.e. fosfomycin
-
?
(S)-2-hydroxypropylphosphonic acid + 2 NADH + O2
cis-(1R,2S)-epoxypropylphosphonic acid + 2 H2O + 2 NAD+
-
i.e. fosfomycin
-
?
(S)-2-hydroxypropylphosphonic acid + 2 NADH + O2
cis-(1R,2S)-epoxypropylphosphonic acid + 2 H2O + 2 NAD+
bidentate mode of substrate binding to the active site iron in the solution state is established. The binding of the hydroxyl group of the substrates determines the site of oxidation. Use of (R)-2-hydroxypropylphosphonic acid yields the 2-keto-adduct rather than the epoxide
i.e. fosfomycin
-
?
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
(R)-2-hydroxypropylphosphonate + H2O2
2-oxopropylphosphonic acid + H2O
-
-
-
?
(R)-2-hydroxypropylphosphonic acid + H2O2
2-oxo-propylphosphonic acid + H2O
-
-
-
?
(S)-2-hydroxypropylphosphonate + H2O2
(1R,2S)-1,2-epoxypropylphosphonate + H2O
-
-
-
?
(S)-2-hydroxypropylphosphonate + H2O2
cis-fosfomycin + trans-fosfomycin + 2 H2O
-
-
-
?
(S)-2-hydroxypropylphosphonic acid + 2 NADH + O2
cis-(1R,2S)-epoxypropylphosphonic acid + 2 H2O + 2 NAD+
-
i.e. fosfomycin
-
?
(R)-2-hydroxypropylphosphonate + H2O2
2-oxo-propylphosphonic acid + H2O
-
-
-
-
?
(R)-2-hydroxypropylphosphonate + NADH + H+ + O2
2-oxopropylphosphonic acid + NAD+ + H2O
-
-
-
-
?
(S)-2-hydroxypropylphosphonate + H2O2
(1R,2S)-1,2-epoxypropylphosphonate + H2O
-
-
-
-
?
(S)-2-hydroxypropylphosphonate + NADH + H+ + O2
(1R,2S)-epoxypropylphosphonate + NAD+ + H2O
-
-
i.e. fosfomycin
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
FAD
FMN and FAD greatly enhance production of fosfomycin. The flavin coenzyme is unlikely an integral part of the epoxidase
FAD
FMN or FAD increase level of fosfomycin production. The effect of FMN is slightly better than that of FAD. The flavin coenzyme is not likely to be an integral part of the epoxidase itself, but it may serve as a surrogate for the putative electron mediator in the in vitro assay
FMN
a straightforward epoxidase mechanism is proposed that depends on the Lewis acid properties of divalent cations and the redox properties of FMN
FMN
FMN and FAD greatly enhance production of fosfomycin. The flavin coenzyme is unlikely an integral part of the epoxidase
FMN
FMN is an artificial electron mediator for the in vitro HppE activity. It accepts a hydride from NADH and then passes electrons on to reduce the iron center of HppE. Several non-flavin electron mediators can replace FMN
FMN
FMN or FAD increase level of fosfomycin production. The effect of FMN is slightly better than that of FAD. The flavin coenzyme is not likely to be an integral part of the epoxidase itself, but it may serve as a surrogate for the putative electron mediator in the in vitro assay
NADH
NADH is a necessary component for (S)-2-hydroxypropylphosphonic acid epoxidation and the overall catalysis is a four-electron redox reaction
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Fe2+
Iron
Zn2+
the recombinant enzyme is active, when reconstituted with Zn2+ or Fe2+
Iron
Fe2+
Fe2+ is the only metal ion that is effective to reconstitute HppE activity. Zn2+ cannot replace Fe2+ in the HppE reaction
Fe2+
the recombinant enzyme is active, when reconstituted with Zn2+ or Fe2+
Iron
mononuclear non-heme iron enzyme. Substrate binds near, and perhaps to, the active site Fe2+ and in doing so organizes the center so that effectively one species is present
Iron
O2 binds to the iron, and substrates bind in a single orientation that strongly perturbs the iron environment. Both functional groups of (S)-2-hydroxypropylphosphonic acid bind to Fe(II) ion at the same time as NO, suggesting that the chelated substrate binding mode dominates in solution. The Fe(II)-substrate chelate structure is important to active fosfomycin formation. This fixed orientation may align the substrate next to the iron-bound activated oxygen species thought to mediate hydrogen atom abstraction from the nearest substrate carbon
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
[(2R)-2-hydroxy-2-(2-methylidenecyclopropyl)ethyl]phosphonate
-
[(2S)-1,1-difluoro-2-hydroxypropyl]phosphonic acid
tight binding inhibitor
[(2R)-2-hydroxy-2-[(1S)-2-methylidenecyclopropyl]ethyl]phosphonate
-
highly effective inhibitor
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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
21400
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
tetramer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
side-chain modification
DOPA formation, probably at Tyr105, is a self-hydroxylation reaction
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystal structures of holoenzyme and the complex with fosfomycin are determined and reveal a two-domain combination and Zn2+ in the active site
hanging drop vapour diffusion method, six X-ray structures of this enzyme: the apoenzyme at 2.0 A resolution, a native Fe2+-bound form at 2.4 A resolution; a tris(hydroxymethyl)aminomethane-Co2+-enzyme complex structure at 1.8 A resolution, a substrate-Co2+-enzyme complex structure at 2.5 A resolution; and two substrate-Fe2+-enzyme complexes at 2.1 and 2.3 A resolution
vapour diffusion method. The hexagonal form displays symmetry consistent with space group P6(1/5)22 and unit-cell parameters a = 86.44, c = 221.56 A, gamma = 120°
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
E142A
complete loss of enzymatic activity in vivo and in vitro, 2.1% of the active site iron as compared to wild-type enzyme, no self-hydroxylation at Tyr105
F182A
the variant favors trans-fosfomycin production from (S)-2-hydroxypropylphosphonate
H138A
inactive in vivo, but in vitro it retains 27% of the active site iron and nearly 20% of the wild-type activity, limited self-hydroxylation (at Tyr105)
H180A
complete loss of enzymatic activity in vivo and in vitro, 5.8% of the active site iron as compared to wild-type enzyme, no self-hydroxylation at Tyr105
Y102F
mutant maintains 60% of the activity of the wild-type enzyme
Y103F
mutant maintains 60% of the activity of the wild-type enzyme
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Cameron, S.; McLuskey, K.; Chamberlayne, R.; Hallyburton, I.; Hunter, W.N.
Initiating a crystallographic analysis of recombinant (S)-2-hydroxypropylphosphonic acid epoxidase from Streptomyces wedmorensis
Acta Crystallogr. Sect. F
61
534-536
2005
Streptomyces wedmorensis (Q56185), Streptomyces wedmorensis
Zhao, Z.; Liu, P.; Murakami, K.; Kuzuyama, T.; Seto, H.; Liu, H.W.
Mechanistic studies of HPP epoxidase: configuration of the substrate governs its enzymatic fate
Angew. Chem. Int. Ed. Engl.
41
4529-4532
2002
Streptomyces wedmorensis (Q56185)
Yan, F.; Li, T.; Lipscomb, J.D.; Liu, A.; Liu, H.W.
Site-directed mutagenesis and spectroscopic studies of the iron-binding site of (S)-2-hydroxypropylphosphonic acid epoxidase
Arch. Biochem. Biophys.
442
82-91
2005
Streptomyces wedmorensis (Q56185), Streptomyces wedmorensis
Liu, P.; Liu, A.; Yan, F.; Wolfe, M.D.; Lipscomb, J.D.; Liu, H.W.
Biochemical and spectroscopic studies on (S)-2-hydroxypropylphosphonic acid epoxidase: a novel mononuclear non-heme iron enzyme
Biochemistry
42
11577-11586
2003
Streptomyces wedmorensis (Q56185)
Yan, F.; Munos, J.W.; Liu, P.; Liu, H.W.
Biosynthesis of fosfomycin, re-examination and re-confirmation of a unique Fe(II)- and NAD(P)H-dependent epoxidation reaction
Biochemistry
45
11473-11481
2006
Streptomyces wedmorensis (Q56185)
Yan, F.; Moon, S.J.; Liu, P.; Zhao, Z.; Lipscomb, J.D.; Liu, A.; Liu, H.W.
Determination of the substrate binding mode to the active site iron of (S)-2-hydroxypropylphosphonic acid epoxidase using 17O-enriched substrates and substrate analogues
Biochemistry
46
12628-12638
2007
Streptomyces wedmorensis (Q56185)
Liu, P.; Murakami, K.; Seki, T.; He, X.; Yeung, S.M.; Kuzuyama, T.; Seto, H.; Liu, H.
Protein purification and function assignment of the epoxidase catalyzing the formation of fosfomycin
J. Am. Chem. Soc.
123
4619-4620
2001
Streptomyces wedmorensis (Q56185)
Liu, P.; Mehn, M.P.; Yan, F.; Zhao, Z.; Que, L. Jr.; Liu, H.W.
Oxygenase activity in the self-hydroxylation of (S)-2-hydroxypropylphosphonic acid epoxidase involved in fosfomycin biosynthesis
J. Am. Chem. Soc.
126
10306-10312
2004
Streptomyces wedmorensis (Q56185), Streptomyces wedmorensis
Hidaka, T.; Goda, M.; Kuzuyama, T.; Takei, N.; Hidaka, M.; Seto, H.
Cloning and nucleotide sequence of fosfomycin biosynthetic genes of Streptomyces wedmorensis
Mol. Gen. Genet.
249
274-280
1995
Streptomyces wedmorensis (Q56185), Streptomyces wedmorensis 144-91 (Q56185)
Higgins, L.J.; Yan, F.; Liu, P.; Liu, H.W.; Drennan, C.L.
Structural insight into antibiotic fosfomycin biosynthesis by a mononuclear iron enzyme
Nature
437
838-844
2005
Streptomyces wedmorensis (Q56185)
McLuskey, K.; Cameron, S.; Hammerschmidt, F.; Hunter, W.N.
Structure and reactivity of hydroxypropylphosphonic acid epoxidase in fosfomycin biosynthesis by a cation- and flavin-dependent mechanism
Proc. Natl. Acad. Sci. USA
102
14221-14226
2005
Streptomyces wedmorensis (Q56185)
Milaczewska, A.; Broclawik, E.; Borowski, T.
On the catalytic mechanism of (S)-2-hydroxypropylphosphonic acid epoxidase (HppE): a hybrid DFT study
Chemistry
19
771-781
2013
Streptomyces wedmorensis (Q56185)
Huang, H.; Chang, W.C.; Pai, P.J.; Romo, A.; Mansoorabadi, S.O.; Russell, D.H.; Liu, H.W.
Evidence for radical-mediated catalysis by HppE: a study using cyclopropyl and methylenecyclopropyl substrate analogues
J. Am. Chem. Soc.
134
16171-16174
2012
Streptomyces wedmorensis (Q56185)
Chang, W.C.; Mansoorabadi, S.O.; Liu, H.W.
Reaction of HppE with substrate analogues: evidence for carbon-phosphorus bond cleavage by a carbocation rearrangement
J. Am. Chem. Soc.
135
8153-8156
2013
Streptomyces wedmorensis (Q56185)
Chang, W.C.; Dey, M.; Liu, P.; Mansoorabadi, S.O.; Moon, S.J.; Zhao, Z.K.; Drennan, C.L.; Liu, H.W.
Mechanistic studies of an unprecedented enzyme-catalysed 1,2-phosphono-migration reaction
Nature
496
114-118
2013
Streptomyces wedmorensis
Huang, H.; Chang, W.C.; Lin, G.M.; Romo, A.; Pai, P.J.; Russell, W.K.; Russell, D.H.; Liu, H.W.
Mechanistic consequences of chiral radical clock probes: analysis of the mononuclear non-heme iron enzyme HppE with 2-hydroxy-3-methylenecyclopropyl radical clock substrates
J. Am. Chem. Soc.
136
2944-2947
2014
Streptomyces wedmorensis
Zhou, S.; Pan, J.; Davis, K.M.; Schaperdoth, I.; Wang, B.; Boal, A.K.; Krebs, C.; Bollinger, J.M.
Steric enforcement of cis-epoxide formation in the radical C-O-coupling reaction by which (S)-2-hydroxypropylphosphonate epoxidase (HppE) produces fosfomycin
J. Am. Chem. Soc.
141
20397-20406
2019
Streptomyces wedmorensis (Q56185)
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