Enzymatic defluorination of fluorinated compounds
- 218 Downloads
Fluorine-containing compounds are widely used because they have properties required in textiles and coatings for electronic, automotive, and outdoor products. However, fluorinated compounds do not easily break down in nature, which has resulted in their accumulation in the environment as well as the human body. Recently, the enzymatic defluorination of fluorine-containing compounds has gained increasing attention. Here, we review the enzymatic defluorination reactions of fluorinated compounds. Furthermore, we review the enzyme engineering strategies for cleaving C–F bonds, which have the highest dissociation energy found in organic compounds.
KeywordsC–F bond Defluorination Fluorine Perfluorinated compound
Because perfluorinated compounds (PFCs) repel both water and oil, they are used as durable repellent treatments for textiles such as outdoor clothes and for home products such as carpets . In addition, PFCs are widely used in the production of fluoropolymers such as polytetrafluoroethylene (trade name, Teflon), which is widely used in coatings for electronic, automotive, and outdoor products .
However, PFCs are harmful to both the natural environment and humans as like other well-known pollutants [1, 3, 4, 5, 6]. Furthermore, PFCs are persistent materials that do not readily break down in natural environments . Some PFCs accumulate in the human body, leading to an increase over time of the residual concentration of PFCs in the blood and organs . For these reasons, the biological decomposition of PFCs has gained increasing attention in recent times. The biological transformation of fluorotelomer alcohols (a type of PFC) was well summarized in several review papers [8, 9]. Nevertheless, the biological pathways underlying PFC decomposition, and the corresponding enzymes, have not been well elucidated.
Here, we review the defluorination reactions of aliphatic and aromatic compounds containing fluorine by native enzymes, including fluoroacetate dehalogenase, fluoroacetate-specific defluorinase, 4-fluorobenzoate dehalogenase, defluorinating enoyl-CoA hydratase/hydrolase, 4-fluorophenol monooxygenase, and peroxygenase-like LmbB2. Furthermore, we review recently developed enzyme engineering strategies for C–F bond cleavage, which give insights into the decomposition of fluorinated compounds. In this review, we do not discuss rarely studied enzymes involved in the biotransformation pathway, such as pyruvate dehydrogenase, maleylacetate reductase, and enol-lactone isomerase [20, 21, 22].
Defluorination by native enzymes
Although the dissociation energy of C–F bond is among the highest found in nature, defluorination of fluoroacetate was identified in microorganisms, such as Burkholderia, Pseudomonas, Delftia, Rhodopseudomonas, and Moraxella [23, 24, 25, 26]. The C–F bond on fluoroacetate can be cleaved by fluoroacetate dehalogenase in such microorganisms . Fluoroacetate dehalogenase functions to the cleavage of C–Cl bond as well as C–F bond. Kinetic parameters (kcat and KM) of Burkholderia fluoroacetate dehalogenase have been known as 9.1 mM and 35 s−1 for fluoroacetate at 30 °C, while 15 mM and 1.5 s−1 for chloroacetate .
In terms of the substrate specificity of fluoroacetate dehalogenase, the importance of Trp150 was asserted by mutation of the amino acid residue . The Trp150Phe mutation resulted in the complete loss of defluorination activity without the lack of dechlorination activity . In a later study using docking simulation and QM/MM calculations, it was suggested that the conformational change for SN2 reaction is favorable in C–F bond rather than C–Cl bond [27, 29]. In the simulation, chloroacetate did not form the reactive conformation for SN2 reaction, because of the longer C–Cl bond . In another study, crystal structures of a Rhodopseudomonas palustris fluoroacetate dehalogenase was determined along the defluorination reaction of fluoroacetate . In such study, Chan and coworkers reported a halide pocket to support three hydrogen bonds which stabilize the fluoride ion in the fluoroacetate dehalogenase-mediated defluorination reaction . Furthermore, the pocket is delicately balanced for fluorine, the smaller halogen atom, for the selectivity of fluoroacetate .
The definition of fluoroacetate-specific defluorinase is still controversial. In a recent study, GSTZ (exactly GSTZ1C) has showed the highest fluoroacetate-specific defluorinase among all GST isozymes, which is just 3% of the total activity of fluoroacetate-specific defluorinase determined in cytosol . Kinetic parameters (Vmax and KM) of fluoroacetate-specific defluorinase activity in rat cytosol have been reported as 27.7 mmol F−/mg protein/h and 3.8 mM for fluoroacetate . In another study, novel fluoroacetate-specific defluorinase, FSD1 was isolated from rat hepatic cytosol . FSD1 showed 81% of the total cytosolic activity of fluoroacetate-specific defluorinase, without GST activity . FSD1 showed about 60% similarity to sorbitol dehydrogenase in amino acid sequence, although defluorination activity of sorbitol dehydrogenase has not been reported .
Enzymes involved in defluorination of fluorinated aromatics
Some enzymes are involved in the degradation pathway of aromatics: 4-fluorobenzoate dehalogenase, defluorinating enoyl-CoA hydratase/hydrolase, 4-fluorophenol monooxygenase, and peroxygenase-like LmbB2 (histidyl-ligated heme enzyme) for the degradation of fluorobenzoate, fluorophenol, fluorobenzene, and fluorotyrosine, respectively.
Enzymes engineering for C–F bond cleavage
The C–F bonds have the highest dissociation energy found in organic compounds, which results in no ready breakdown of the substance containing the C–F bond in natural environments. For the reason, some PFCs had been considered as a serious organic pollutant in the Stockholm convention in 2006. Wide use of F-containing compounds has resulted in the widespread pollution in the world, which has been urging us to develop defluorination technologies for decompose the compounds. As described above, the enzymatic defluorination reactions of fluorinated compounds can be catalyzed by employing native and artificially designed enzymes. It is expected that more successful examples of C-F cleavage will appear through the enzyme engineering integrated with synthetic biology techniques. In the near future, native and artificial defluorination enzymes will play a major role in the reduction of the widely spread pollutant containing C–F bond.
This study was carried out with the support of “Cooperative Research Program for Agricultural Science & Technology Development (Project No. PJ013321022019)”, Rural Development Administration, Republic of Korea. YSJ was supported by a grant from the Ministry of Science and ICT (MSIT) through the National Research Foundation (NRF) of Korea (NRF-2019R1A4A1029125).
YSJ and JHK designed the project. HJS, SWK, and YSJ wrote the manuscript. JHK and DCS revised the manuscript. All authors read and approved the final manuscript.
This study was carried out with the support of “Cooperative Research Program for Agricultural Science & Technology Development (Project No. PJ013321022019)”, Rural Development Administration, Republic of Korea. YSJ was supported by a Grant from the Ministry of Science and ICT (MSIT) through the National Research Foundation (NRF) of Korea (NRF-2019R1A4A1029125).
The authors declare that they have no competing interests.
- 24.Kawasaki H, Yahara H, Tonomura K (1984) Cloning and expression in Escherichia coli of the haloacetate dehalogenase genes from moraxella plasmid pUO1. Agric Biol Chem 48:2627–2632Google Scholar
- 27.Nakayama T, Kamachi T, Jitsumori K, Omi R, Hirotsu K, Esaki N, Kurihara T, Yoshizawa K (2012) Substrate specificity of fluoroacetate dehalogenase: an insight from crystallographic analysis, fluorescence spectroscopy, and theoretical computations. Chem Eur J 18:8392–8402PubMedCrossRefPubMedCentralGoogle Scholar
- 35.Tu LQ, Wright PFA, Rix CJ, Ahokas JT (2006) Is fluoroacetate-specific defluorinase a glutathione S-transferase? Comp Biochem Phys C 143:59–66Google Scholar
Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.