On the Transformation of (S)-2-Hydroxypropylphosphonic Acid into Fosfomycin in Streptomyces fradiae—A Unique Method of Epoxide Ring Formation

Woschek, Anna; Wuggenig, Frank; Peti, Wolfgang; Hammerschmidt, Friedrich

doi:10.1002/1439-7633(20020902)3:9<829::aid-cbic829>3.0.co;2-v

Cited by 16 publications

(16 citation statements)

References 14 publications

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“…This gives the major product fosfomycin by a mechanism, consistent with previous findings (7). A very minor product (not depicted) is the trans-epoxide, which would be generated after rotation (180°) about the C1OC2 bond (7). The structural differences between HPPE-Zn and HPPE-Fos and the stereochemistry of the reaction are consistent with the hypothesis that FMN would bind over toward Tyr-103, Arg-97, Asn-99, and the flexible loop discussed earlier.…”

Section: Resultssupporting

confidence: 92%

“…The O2 alkoxide attacks C1 from the opposite side of H* abstraction and forms the C1OO2 bond with inversion of configuration. This gives the major product fosfomycin by a mechanism, consistent with previous findings (7). A very minor product (not depicted) is the trans-epoxide, which would be generated after rotation (180°) about the C1OC2 bond (7).…”

Section: Resultssupporting

confidence: 84%

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Structure and reactivity of hydroxypropylphosphonic acid epoxidase in fosfomycin biosynthesis by a cation- and flavin-dependent mechanism

McLuskey

Cameron

Hammerschmidt

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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The biosynthesis of fosfomycin, an oxirane antibiotic in clinical use, involves a unique epoxidation catalyzed by (S)-2-hydroxypropylphosphonic acid epoxidase (HPPE). The reaction is essentially dehydrogenation of a secondary alcohol. A high-resolution crystallographic analysis reveals that the HPPE subunit displays a two-domain combination. The C-terminal or catalytic domain has the cupin fold that binds a divalent cation, whereas the N-terminal domain carries a helix-turn-helix motif with putative DNA-binding helices positioned 34 Å apart. The structure of HPPE serves as a model for numerous proteins, of ill-defined function, predicted to be transcription factors but carrying a cupin domain at the C terminus. Structure-reactivity analyses reveal conformational changes near the catalytic center driven by the presence or absence of ligand, that HPPE is a Zn 2؉ ͞Fe 2؉ -dependent epoxidase, proof that flavin mononucleotide is required for catalysis, and allow us to propose a simple mechanism that is compatible with previous experimental data. The participation of the redox inert Zn 2؉ in the mechanism is surprising and indicates that Lewis acid properties of the metal ions are sufficient to polarize the substrate and, aided by flavin mononucleotide reduction, facilitate the epoxidation.enzyme ͉ epoxidation ͉ iron ͉ zinc

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Section: Resultssupporting

confidence: 92%

Section: Resultssupporting

confidence: 84%

Structure and reactivity of hydroxypropylphosphonic acid epoxidase in fosfomycin biosynthesis by a cation- and flavin-dependent mechanism

McLuskey

Cameron

Hammerschmidt

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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“…It has previously been demonstrated that the C-1 pro- R hydrogen atom is abstracted in the conversion of ( S )-HPP to fosfomycin (25, 38); so (1 R ,2 S )-1-[ 2 H]-HPP ( 9 ) was used as the labeled substrate to determine the [ 2 H]-KIE. For the turnover of ( R )-HPP to 8 , it is the C-2 hydrogen atom that is abstracted (37), so that the [ 2 H]-KIE was measured using ( R )-2-[ 2 H]-HPP ( 15 ).…”

Section: Resultsmentioning

confidence: 99%

Purification and Characterization of the Epoxidase Catalyzing the Formation of Fosfomycin from Pseudomonas syringae

et al. 2008

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The final step in the biosynthesis of fosfomycin in Streptomyces wedmorensis is catalyzed by (S)-2-hydroxypropylphosphonic acid (HPP) epoxidase (Sw-HppE). A homologous enzyme from Pseudomonas syringae has recently been isolated whose encoding gene (orf3) shares relatively low sequence homology to the corresponding Sw-HppE gene. This purified P. syringae protein was determined to catalyze the epoxidation of (S)-HPP to fosfomycin and the oxidation of (R)-HPP to 2-oxopropylphosphonic acid under the same conditions as Sw-HppE. Therefore, this protein is indeed a true HPP epoxidase and is termed Ps-HppE. Like Sw-HppE, Ps-HppE was determined to be posttranslationally modified by the hydroxylation of a putative active site tyrosine (Tyr95). Analysis of the Fe(II)-center by EPR spectroscopy using NO as a spin probe and molecular oxygen surrogate reveals that Ps-HppE's metal center is similar, but not identical, to that of Sw-HppE. The identity of the rate determining step for the (S)-HPP and (R)-HPP reactions was determined by measuring primary deuterium kinetic effects, and the outcome of these results were correlated with density functional theory calculations. Interestingly, the reaction using the non-physiological substrate (R)-HPP was 1.9 times faster than that with (S)-HPP for both Ps-HppE and Sw-HppE. This is likely due to the difference in bond dissociation energy of the abstracted hydrogen atom for each respective reaction. Thus, despite low amino acid sequence identity, Ps-HppE is a close mimic of Sw-HppE, representing a second example of a non-heme iron-dependent enzyme capable of catalyzing dehydrogenation of a secondary alcohol to form a new C-O bond.Fosfomycin (1) is a clinically useful antibiotic (1) for the treatment of limb-threatening diabetic foot infections (2) and lower urinary tract infections. It has been shown to be effective against ciprofloxacin-resistant Escherichia coli (3), as well as methicillin-resistant (4) and vancomycin-resistant (5) strains of Staphylococcus aureus. The antimicrobial activity of fosfomycin is due to the inactivation of UDP-GlcNAc-3-O-enolpyruvyltransferase (MurA), which catalyzes the first committed step in the biosynthesis of peptidoglycan, the main component of the bacterial cell wall (6,7). NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptFosfomycin belongs to a steadily growing family of natural products containing a C-P bond (8) (9). Members of this family, which include fosmydomicin (10) and bialaphos (11) are all derived from phosphoenolpyruvate (PEP, 2). The C-P bonds in these compounds are formed through an intramolecular rearrangement reaction catalyzed by PEP mutase, resulting in the conversion of PEP (2) to phosphopyruvate (PnPy, 3) ( Fig. 1) (12)(13)(14). Since the equilibrium between PEP and PnPy favors PEP, the decarboxylation catalyzed by the second enzyme in the biosynthetic pathway, PnPy decarboxylase, provides the driving force to shift the equilibrium toward C-P bond formation (15). The next two steps in the pathway...

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“…The stereochemical course of this reaction has been investigated through the use of stereospecifically deuterated HPP. Findings from these studies show that only the pro-(R) hydrogen at C1 is abstracted [107]. The same researchers also report the isolation of small amounts of the trans isomer of fosfomycin from the culture broth [108].…”

Section: Fosfomycinmentioning

confidence: 99%

“…The same researchers also report the isolation of small amounts of the trans isomer of fosfomycin from the culture broth [108]. This compound could be formed by rotation around the C1-C2 bond at the C1-radical stage [107]; a tentative reaction cycle is shown in Scheme 10.27. Interestingly, HppE is capable of oxidizing both the native substrate (S)-HPP and also its enantiomer (R)-HPP.…”

Section: Fosfomycinmentioning

confidence: 99%

The Biosynthesis of Epoxides

Grüschow

Sherman

2006

Aziridines and Epoxides in Organic Synthesis

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On the Transformation of (S)-2-Hydroxypropylphosphonic Acid into Fosfomycin in Streptomyces fradiae—A Unique Method of Epoxide Ring Formation

Cited by 16 publications

References 14 publications

Structure and reactivity of hydroxypropylphosphonic acid epoxidase in fosfomycin biosynthesis by a cation- and flavin-dependent mechanism

Structure and reactivity of hydroxypropylphosphonic acid epoxidase in fosfomycin biosynthesis by a cation- and flavin-dependent mechanism

Purification and Characterization of the Epoxidase Catalyzing the Formation of Fosfomycin from Pseudomonas syringae

The Biosynthesis of Epoxides

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