Enzyme-catalyzed C–F bond formation and cleavage
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Organofluorines are widely used in a variety of applications, ranging from pharmaceuticals to pesticides and advanced materials. The widespread use of organofluorines also leads to its accumulation in the environment, and two major questions arise: how to synthesize and how to degrade this type of compound effectively? In contrast to a considerable number of easy-access chemical methods, milder and more effective enzymatic methods remain to be developed. In this review, we present recent progress on enzyme-catalyzed C–F bond formation and cleavage, focused on describing C–F bond formation enabled by fluorinase and C–F bond cleavage catalyzed by oxidase, reductase, deaminase, and dehalogenase.
KeywordsOrganofluorines C–F bonds Enzyme-catalyzed Degradation Formation
- class I BCR
catalyzed by class I BzCoA reductase
Incorporation of fluorine into organic compounds usually endows organofluorines with unique chemical and physical properties, a strategy that has been successfully applied in agrochemicals, materials science, and pharmaceutical chemistry (Phelps 2004; Müller et al. 2007; Shah and Westwell 2007; Hagmann 2008; Nenajdenko et al. 2015; Zhang et al. 2016; Lowe et al. 2017). Especially in medicinal chemistry, the unique elemental properties of fluorine have been proved to enhance metabolic stability and alter pharmacokinetic characteristics without increasing the apparent spatial volume; thus, more than 20% of drugs are organofluorines (Zhou et al. 2016; Gillis et al. 2015; Spooner et al. 2019). The wide application of organofluorines has motivated fast methodology development for fluorine incorporation (Purser et al. 2008; Berger et al. 2011). In contrast to their synthesis, their degradation has also attracted significant attention due to their increased use and the cumulative pollution resulting from their high stabilities.
Chemists have developed versatile methods for the formation and cleavage of C–F bonds, but these methods usually require harsh conditions and are not environment friendly (Dillert et al. 2007; Lin et al. 2012; Sulbaek Andersen et al. 2005). To solve these problems, development of mild and green methods is urgently needed. Biocatalysis has been playing an increasingly more important role in modern chemistry due to its high efficiency, specific selectivity, and more environmentally friendly characteristics compared to chemical catalysis. Thus, introducing biocatalytic methods into organic fluorine chemistry is a good choice to counter the deficiencies of chemical catalysis (Kim et al. 2000; Liu and Avendaño 2013; Murphy 2016; Rotander et al. 2012). Although biocatalysis has achieved significant progress in recent years, the field of biocatalytic C–F bond formation and cleavage is almost at an open stage.
Since fluorine atoms are very small and strongly electro-negative, when in an aqueous system fluoride ions are always tightly wrapped by the water molecules, and thus, it is very difficult to form C–F bonds in an aqueous system (O’Hagan 2008; Ni and Hu 2016). Therefore, fluorine-containing natural products are very rare despite the fact that elemental fluorine is the most fecund halogen in the Earth’s crust (O’Hagan and Deng 2014). To the best of our knowledge, only two different examples of enzyme-catalyzed C–F bond formation have been reported: one is catalyzed by a mutant of glycosyltransferase, which catalyzes α-fluoroglycosides as transient intermediates from DNP-activated sugars (Zechel et al. 2001), and the other is the natural fluorinase, prompting the conversion of 5′-fluoro-5′-deoxyribose-1-phosphate (5′-FRP) from S-adenosyl-l-methionine (SAM) (Deng et al. 2004). In this review, we focus on summarizing the recent progress in mining and directed evolution of fluorinase and expect to inspire the development of more unnatural fluorinases in the future.
The C–F bond is the strongest σ bond, and thus, it is difficult to cleave it under mild conditions (Goldman 1969; Lemal 2004). When hydrogen is substituted by fluorine, the metabolic stability of the compounds will be significantly improved; this property benefits the pharmaceutical industry, but leads to the accumulation of organofluorines in the environment (Wang et al. 2016). Enzyme-catalyzed C–F bond cleavage has attracted attention from researchers in environmental protection, C–F bond activation, and enzymology, and several reviews have been published on the subject (Natarajan et al. 2005). However, in recent years, reports in this area have been very rare. Herein, therefore, we present recently published examples of enzyme-catalyzed C–F bond cleavage, dividing them into two types: hydrolytic defluorination, and oxireductive defluorination. Hopefully, this review will attract increasing numbers of workers to this important field.
Enzyme-catalyzed C–F bond formation
Comparative kinetic data of known fluorinase enzymes
Fluorinase (FIA) source
SAM Km (μM)
Turnover no. kcat (min−1)
Specificity constant kcat/Km (mM−1 min−1)
29.2 ± 2.41
Schaffrath et al. 2003
Streptomyce sp. MA37
82.4 ± 18.6
Deng et al. 2014
27.8 ± 4.23
Wang et al. 2014
Actinoplanes sp. N902-109
45.8 ± 7.91
Deng et al. 2014
RCC (analytical) (%)
RCCa (overall) (%)
8 ± 1
7 ± 1
11 ± 2
8 ± 2
32 ± 3
24 ± 2
fah2114 (F213Y, A279L)
46 ± 2
34 ± 3
Enzyme-catalyzed C–F bond cleavage
Large-scale applications of fluorinated compounds have caused increasing environmental concerns due to their toxicity, global warming potential, environmental persistence, and bioaccumulation character (Douvris and Ozerov 2008; Houde et al. 2006). Environmental biotransformation, one of the most promising strategies with the lowest energy consumption, has provided some encouraging results in cleaving the highly stable C–F bond, the dissociation energy of which is the highest among all the natural products. At present, there are two ways of catalyzing the cleavage of C–F bonds by enzymes: hydrolytic defluorination and oxireductive defluorination.
Organofluorines play an increasingly important role in the pharmaceutical and agrochemical industries, making the prospect of using enzymatic reactions to form C–F bonds bright. However, the extensive use of organofluorines has also caused environmental pollution, and thus, development of a mild green enzyme to degrade these compounds is a matter of urgency. With its highly catalytic selectivity and environmental friendliness, enzymatic catalysis will play an increasingly more important role in fluorine organic chemistry. In this review, details of the formation of C–F bonds catalyzed by fluorinase and the cleavage of C–F bonds by oxidase, reductase deaminase, and fluoroacetate dehalogenase are demonstrated. These fluorinase and defluorinase all have been isolated and identified for more than a decade, and their catalytic mechanisms illuminated. However, a narrow substrate range or low activity has hindered their application. With the fast development of biotechnology, mining new enzymes or improving their properties by directed evolution holds promise to eliminate these barriers, which will greatly accelerate the development of enzymatic organic fluorine chemistry.
J.-b.W. thanks and the support from Huxiang High-level Talent Gathering Project of Hunan Province (2019RS1040) and the start-up funding from Hunan Normal University.
JW and WT conceived and wrote this paper. QH and ML were involved collecting related material and critical reading of this paper. All authors read and approved the final manuscript.
Ethics approval and consent to participate
Consent for publication
The authors declare that they have no competing interests.
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