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ABSTRACT: 5'-Fluoro-5'-deoxy-D-ribose-1-phosphate (FDRP) is identified as a biosynthetic intermediate during fluorometabolite biosynthesis in Streptomyces cattleya.
Chemical Communications 04/2004; · 6.17 Impact Factor
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ABSTRACT: The energies of the gauche and anti conformers of 2-fluoroethylamine, 2-fluoroethanol and their protonated analogues are calculated using density functional theory. Unlike the non protonated systems, the protonated systems show a strong gauche effect where the C-F and the C-(+)NH(3) or C-F and C-(+)OH(2) bonds are gauche rather than anti to each other. Single crystal X-ray diffraction studies of 2-fluoroethylammonium compounds identify the same conformational preference.
Organic & Biomolecular Chemistry 04/2004; 2(5):732-40. · 3.70 Impact Factor
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ABSTRACT: Fluorine is the thirteenth most abundant element in the earth's crust, but fluoride concentrations in surface water are low and fluorinated metabolites are extremely rare. The fluoride ion is a potent nucleophile in its desolvated state, but is tightly hydrated in water and effectively inert. Low availability and a lack of chemical reactivity have largely excluded fluoride from biochemistry: in particular, fluorine's high redox potential precludes the haloperoxidase-type mechanism used in the metabolic incorporation of chloride and bromide ions. But fluorinated chemicals are growing in industrial importance, with applications in pharmaceuticals, agrochemicals and materials products. Reactive fluorination reagents requiring specialist process technologies are needed in industry and, although biological catalysts for these processes are highly sought after, only one enzyme that can convert fluoride to organic fluorine has been described. Streptomyces cattleya can form carbon-fluorine bonds and must therefore have evolved an enzyme able to overcome the chemical challenges of using aqueous fluoride. Here we report the sequence and three-dimensional structure of the first native fluorination enzyme, 5'-fluoro-5'-deoxyadenosine synthase, from this organism. Both substrate and products have been observed bound to the enzyme, enabling us to propose a nucleophilic substitution mechanism for this biological fluorination reaction.
Nature 03/2004; 427(6974):561-5. · 36.28 Impact Factor
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ABSTRACT: Fluorine is the thirteenth most abundant element in the earth's crust, but fluoride concentrations in surface water are low and fluorinated metabolites are extremely rare
Nature 02/2004; 427(6974):561-565. · 36.28 Impact Factor
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ABSTRACT: Organofluorine compounds are widely prepared throughout the chemicals industry, but their prepararion generally requires harsh fluorinating reagents and non-aqueous solvents. On the other hand, biology has hardly exploited organofluorine compounds. A very few organisms synthesize organofluorine metabolites, suggesting they have evolved a mechanism to overcome the kinetic desolvation barrier to utilizing F(-)(aq). Here, the purification and crystallization of an enzyme from Streptomyces cattleya which is responsible for the synthesis of the C-F bond during fluoroacetate and 4-fluorothreonine biosynthesis is reported. The protein crystallizes in space group C222(1), with unit-cell parameters a = 75.9, b = 130.3, c = 183.4 A, alpha = beta = gamma = 90 degrees. Data were recorded to 1.9 A at the ESRF. The structure of the protein should provide important insights into the biochemical process of C-F bond formation.
Acta Crystallographica Section D Biological Crystallography 01/2004; 59(Pt 12):2292-3. · 12.62 Impact Factor
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ABSTRACT: The use of the key enzyme involved in carbon–fluorine bond formation in Streptomyces cattleya catalysing the formation of 5′-fluoro-5′-deoxyadenosine (5′-FDA) from fluoride ion and S-adenosyl-l-methionine (SAM) was explored for its potential application in fluorine-18 labelling of the adenosine derivative. Enzymatic radiolabelling of [18F]-5′-FDA was successfully carried out starting from SAM and [18F]HF when the concentration of the enzyme preparation was increased from sub-mg/ml values to mg/ml values. The purity of the enzyme had no measurable effect on the radiochemical yield of the reaction and the radiochemical purity of [18F]-5′-FDA. Copyright © 2003 John Wiley & Sons, Ltd.
Journal of Labelled Compounds 10/2003; 46(13):1181 - 1189.
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ABSTRACT: 5'-fluorodeoxyadenosine synthase, a C-F bond-forming enzyme, has been purified from Streptomyces cattleya. The enzyme mediates a reaction between inorganic fluoride and S-adenosyl-L-methionine (SAM) to generate 5'-fluoro-5'-deoxyadenosine. The molecular weight of the monomeric protein is shown to be 32.2 kDa by electrospray mass spectrometry. The kinetic parameters for SAM (K(m) 0.42 mM, V(max) 1.28 U/mg) and fluoride ion (K(m) 8.56 mM, V(max) 1.59 U/mg) have been evaluated. Both S-adenosyl-L-homocysteine (SAH) and sinefungin were explored as inhibitors of the enzyme. SAH emerged as a potent competitive inhibitor (K(i) 29 microM) whereas sinefungin was only weakly inhibitory.
FEBS Letters 08/2003; 547(1-3):111-4. · 3.54 Impact Factor
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ABSTRACT: Organofluorine compounds are rare in Nature, with only a handful known to be produced by some species of plant and two microorganisms. Consequently, the mechanism of enzymatic carbon-fluorine bond formation is poorly understood. The bacterium Streptomyces cattleya biosynthesises fluoroacetate and 4-fluorothreonine as secondary metabolites and is a convenient system to study the biosynthesis and enzymology of fluorometabolite production. Using stable-isotope labelled precursors it has been shown that there is a common intermediate in the biosynthesis of the fluorometabolites, which has recently been identified as fluoroacetaldehyde. Studies with cell-free extracts of S. cattleya have identified two enzymes, an aldehyde dehydrogenase and a threonine transaldolase, that are involved in the biotransformation of fluoroacetaldehyde to fluoroacetate and 4-fluorothreonine.
Chemosphere 08/2003; 52(2):455-61. · 3.21 Impact Factor
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ABSTRACT: A sensitive method for the configurational analysis of (R)- and (S)-[2H1]-fluoroacetate has been developed using 2H[1H]-NMR in a chiral liquid crystalline solvent. This has enabled biosynthetic experiments to be conducted which reveal stereochemical details on biological fluorination occurring during the biosynthesis of fluoroacetate and 4-fluorothreonine in the bacterium Streptomyces cattleya. In particular, feeding experiments to S. cattleya with isotopically labeled (1R, 2R)- and (1S, 2R)-[1-2H1]-glycerol 3d and 3e and [2,3-2H(4)]-succinate 4a gave rise to samples of enantiomerically enriched [2-2H1]-fluoroacetates 1a. The predominant enantiomer resulting from each experiment suggests that the stereochemical course of biological fluorination takes place with an overall retention of configuration between a glycolytic intermediate and fluoroacetate 1. Consequently, this outcome suggests that the stereochemical course of the recently identified fluorinase enzyme which mediates a reaction between fluoride ion and S-adenosyl-l-methionine (SAM), occurs with an inversion of configuration.
Journal of the American Chemical Society 02/2003; 125(2):379-87. · 9.91 Impact Factor
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Angewandte Chemie International Edition 11/2002; 41(20):3913-5. · 13.45 Impact Factor
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ABSTRACT: The conversion of littorine to hyoscyamine has been investigated by feeding deuterium labelled (RS)-[2-(2)H]-, [3, 3-(2)H(2)]-, [2, 3, 3-(2)H(3)]- phenyllactic acids to transformed root cultures of Datura stramonium. Isolation and GC-MS analyses of the isotope incorporation into the resultant hyoscyamine does not support the involvement of a vicinal interchange process operating during the isomerisation of littorine to hyoscyamine. Additionally a metabolism study with [1'-13C, 3', 3'-(2)H(2)]-hyoscyamine has established that the alkaloid is metabolically stable at C-3' with no evidence for a reversible in vivo oxidation process to the corresponding aldehyde. The data do not support an S-adenosy-L-methionine (SAM 5)/co-enzyme-B(12) mediated process for the isomerisation of littorine to hyoscyamine.
Phytochemistry 11/2002; 61(3):323-9. · 3.35 Impact Factor
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ABSTRACT: Lipase-catalysed (trans)esterification reactions in homogenous perfluorocarbon-hydrocarbon solvents enabled direct enantiomeric partitioning (up to 95% ee) of the products by liquid-liquid separation.
Chemical Communications 09/2002; · 6.17 Impact Factor
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ABSTRACT: (+/-)-Erythro- and (+/-)-threo-9,10-difluorostearic acids, which differ only by a stereogenic interconversion of a single C-F bond, have significantly different conformational stabilities.
Chemical Communications 07/2002; · 6.17 Impact Factor
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ABSTRACT: The enantiomeric assay of fluoroalkanes using 2H-NMR in a chiral liquid crystalline medium is demonstrated, and at its limit the enantiomers of [5-(2)H]-5-fluorodecane were successfully resolved.
Chemical Communications 05/2002; · 6.17 Impact Factor
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ABSTRACT: Although fluorine in the form of fluoride minerals is the most abundant halogen in the Earth's crust, only 12 naturally occurring organofluorine compounds have so far been found, and how these are biosynthesized remains a mystery. Here we describe an enzymatic reaction that occurs in the bacterium Streptomyces cattleya and which catalyses the conversion of fluoride ion and S-adenosylmethionine (SAM) to 5'-fluoro-5'-deoxyfluoroadenosine (5'-FDA). To our knowledge, this is the first fluorinase enzyme to be identified, a discovery that opens up a new biotechnological opportunity for the preparation of organofluorine compounds.
Nature 04/2002; 416(6878):279. · 36.28 Impact Factor
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ABSTRACT: A synthetic route to isotopically labelled forms of the tropane alkaloid hyoscyamine, including (RS)-[3′, 3′,-2H2]- (2a) and (RS)-[1′-13C, 3′, 3′,-2H2]- (2b) hyoscyamines, involving the reaction between phenylacetyl tropine and formaldehyde is described. The isotopically labelled products enable the metabolism of hyoscyamine to be studied in plants such as Datura stramonium. Copyright © 2002 John Wiley & Sons, Ltd.
Journal of Labelled Compounds 01/2002; 45(3):191 - 198.
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Angewandte Chemie International Edition 01/2002; 40(23):4479-4481. · 13.45 Impact Factor
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Angewandte Chemie International Edition 11/2001; 40(23):4479 - 4481. · 13.45 Impact Factor
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J. Chem. Soc., Perkin Trans. 2. 01/2000;
