Biology:Fluorinase
| Fluorinase (adenosyl-fluoride synthase) | |||||||||
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| Identifiers | |||||||||
| EC number | 2.5.1.63 | ||||||||
| Databases | |||||||||
| IntEnz | IntEnz view | ||||||||
| BRENDA | BRENDA entry | ||||||||
| ExPASy | NiceZyme view | ||||||||
| KEGG | KEGG entry | ||||||||
| MetaCyc | metabolic pathway | ||||||||
| PRIAM | profile | ||||||||
| PDB structures | RCSB PDB PDBe PDBsum | ||||||||
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The fluorinase enzyme (EC 2.5.1.63, also known as adenosyl-fluoride synthase) catalyzes the reaction between fluoride ion and the co-factor S-adenosyl-L-methionine to generate L-methionine and 5'-fluoro-5'-deoxyadenosine, the first committed product of the fluorometabolite biosynthesis pathway.[1] The fluorinase was originally isolated from the soil bacterium Streptomyces cattleya, but homologues have since been identified in a number of other bacterial species, including Streptomyces sp. MA37, Nocardia brasiliensis and Actinoplanes sp. N902-109.[2] This is the only known enzyme capable of catalysing the formation of a carbon-fluorine bond, the strongest single bond in organic chemistry.[3]
A homologous chlorinase enzyme, which catalyses the same reaction with chloride rather than fluoride ion, has been isolated from Salinospora tropica, from the biosynthetic pathway of salinosporamide A.[4]
Reactivity
The fluorinase catalyses an SN2-type nucleophilic substitution at the C-5' position of SAM, while L-methionine acts as a neutral leaving group.[5][6] The fluorinase-catalysed reaction is estimated to be between 106[6] and 1015[7] times faster than the uncatalysed reaction, a significant rate enhancement. Despite this, the fluorinase is still regarded as a slow enzyme, with a turnover number (kcat) of 0.06 min−1.[8] The high kinetic barrier to reaction is attributed to the strong solvation of fluoride ion in water, resulting in a high activation energy associated with stripping solvating water molecules from aqueous fluoride ion, converting fluoride into a potent nucleophile within the active site.
The reaction catalysed by the fluorinase is reversible, and upon incubation of 5'-fluoro-5'-deoxyadenosine and L-methionine with the fluorinase, SAM and fluoride ion are produced.[9] Replacing L-methionine with L-selenomethionine results in a 6-fold rate enhancement of the reverse reaction,[9] due to the increased nucleophilicity of the selenium centre compared to the sulfur centre.
The fluorinase shows a degree of substrate tolerance for halide ion, and can also use chloride ion in place of fluoride ion. While the equilibrium for reaction between SAM and fluoride ion lies towards products FDA and L-methionine, the equilibrium position is reversed in the case for chloride ion. Incubation of SAM and chloride ion with the fluorinase does not result in generation of 5'-chloro-5'-deoxyadenosine (ClDA), unless an additional enzyme, an L-amino acid oxidase, is added. The amino acid oxidase removes the L-methionine from the reaction, converting it to the corresponding oxo-acid.
The halide preference, coupled to the position of the two reaction equilibria allows for a nett transhalogenation reaction to be catalysed by the enzyme.[9] Incubation of 5'-chloro nucleosides with the enzyme, along with catalytic L-selenomethionine or L-methionine results in the production of 5-fluoro nucleosides. When [18F]fluoride is used, this transhalogenation reaction can be used for the synthesis of radiotracers for positron emission tomography.[10][11]
Structural studies
As of late 2007, 9 structures have been solved for this class of enzymes, with PDB accession codes 1RQP, 1RQR, 2C2W, 2C4T, 2C4U, 2C5B, 2C5H, 2CBX, and 2CC2.
The names given to the enzyme come not from the structure, but from the function: 5-Fluoro-5-deoxyadenosine is the molecule synthesised. The structure is homologous to the duf-62 enzyme series. The enzyme is a dimer of trimers (2 molecules each with three subunits). The active sites are located between these subunits (subunit interfaces), each can bind to one SAM molecule at a time.[12]
Fluorometabolite biosynthesis
See also
References
- ↑ 1.0 1.1 "Biochemistry: biosynthesis of an organofluorine molecule". Nature 416 (6878): 279. March 2002. doi:10.1038/416279a. PMID 11907567.
- ↑ "Identification of fluorinases from Streptomyces sp MA37, Norcardia brasiliensis, and Actinoplanes sp N902-109 by genome mining". ChemBioChem 15 (3): 364–8. February 2014. doi:10.1002/cbic.201300732. PMID 24449539.
- ↑ "Understanding organofluorine chemistry. An introduction to the C-F bond". Chemical Society Reviews 37 (2): 308–19. February 2008. doi:10.1039/b711844a. PMID 18197347.
- ↑ "Discovery and characterization of a marine bacterial SAM-dependent chlorinase". Nature Chemical Biology 4 (1): 69–74. January 2008. doi:10.1038/nchembio.2007.56. PMID 18059261.
- ↑ "Enzymatic fluorination in Streptomyces cattleya takes place with an inversion of configuration consistent with an SN2 reaction mechanism". ChemBioChem 5 (5): 685–90. May 2004. doi:10.1002/cbic.200300839. PMID 15122641.
- ↑ 6.0 6.1 "Insight into enzymatic C-F bond formation from QM and QM/MM calculations". Journal of the American Chemical Society 127 (39): 13643–55. October 2005. doi:10.1021/ja053875s. PMID 16190730.
- ↑ "Catalysis by desolvation: the catalytic prowess of SAM-dependent halide-alkylating enzymes". Journal of the American Chemical Society 135 (39): 14473–5. October 2013. doi:10.1021/ja406381b. PMID 24041082.
- ↑ "Mechanism of enzymatic fluorination in Streptomyces cattleya". Journal of the American Chemical Society 129 (47): 14597–604. November 2007. doi:10.1021/ja0731569. PMID 17985882.
- ↑ 9.0 9.1 9.2 "The fluorinase from Streptomyces cattleya is also a chlorinase". Angewandte Chemie 45 (5): 759–62. January 2006. doi:10.1002/anie.200503582. PMID 16370017.
- ↑ "Fluorinase mediated C-(18)F bond formation, an enzymatic tool for PET labelling". Chemical Communications (6): 652–4. February 2006. doi:10.1039/b516861a. PMID 16446840.
- ↑ "A two-step fluorinase enzyme mediated (18)F labelling of an RGD peptide for positron emission tomography". Chemical Communications 51 (70): 13542–5. September 2015. doi:10.1039/c5cc05013h. PMID 26221637.
- ↑ "Crystal structure and mechanism of a bacterial fluorinating enzyme". Nature 427 (6974): 561–5. February 2004. doi:10.1038/nature02280. PMID 14765200.
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