Development of a Hydrolysis-Based Small-Molecule Hydrogen Selenide (H Se) Donor† Cite This: Chem
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Chemical Science View Article Online EDGE ARTICLE View Journal | View Issue Development of a hydrolysis-based small-molecule hydrogen selenide (H Se) donor† Cite this: Chem. Sci.,2019,10,10723 2 All publication charges for this article Turner D. Newton and Michael D. Pluth * have been paid for by the Royal Society of Chemistry Selenium is essential to human physiology and has recently shown potential in the treatment of common pathophysiological conditions ranging from arsenic poisoning to cancer. Although the precise metabolic and chemical pathways of selenium incorporation into biomolecules remain somewhat unclear, many À such pathways proceed through hydrogen selenide (H2Se/HSe ) formation. Despite this importance, well-characterized chemistry that enables H2Se release under controlled conditions remains lacking. Motivated by this need, we report here the development of a hydrolysis-based H2Se donor (TDN1042). 31 77 Utilizing P and Se NMR experiments, we demonstrate the pH dependence of H2Se release and characterize observed reaction intermediates during the hydrolysis mechanism. Finally, we confirm H2Se Received 12th September 2019 release using electrophilic trapping reagents, which not only demonstrates the fidelity of this donor Accepted 11th October 2019 platform but also provides an efficient method for investigating future H2Se donor motifs. Taken Creative Commons Attribution-NonCommercial 3.0 Unported Licence. DOI: 10.1039/c9sc04616j together, this work provides an early example of an H2Se donor that functions through a well-defined rsc.li/chemical-science and characterized mechanism. Introduction decient environments oen result in the preferential expres- sion of these proteins, whereas selenium-rich environments Selenium is oen regarded as a toxic metalloid, but it is also an result in the upregulation of selenium excretion pathways to 8 essential bioinorganic dietary micronutrient.1–3 For example, mitigate selenium toxicity. Many of these pathways are geographic regions with selenium-decient soil display hypothesized to proceed through the intermediate formation of This article is licensed under a À unusually high occurrences of conditions including Keshan and hydrogen selenide (H2Se/HSe ), which is an important yet 9 Kashin-Beck diseases in the population, which are both tied to elusive small biomolecule of interest (Fig. 1). a dietary scarcity of selenium.4,5 Dietary sources of selenium are A common approach to increase the bioavailability of sele- À Open Access Article. Published on 11 October 2019. Downloaded 9/28/2021 8:08:42 AM. 2 typically selenomethionine (SeMet) and selenate salts (SeO4 ), nium is to use exogenous synthetic selenium-containing small which must pass through complex metabolic pathways prior to molecules. For example, the organoselenium compound ebse- incorporation into selenium-containing biomolecules.6 In the len mimics the behavior of glutathione peroxidase and exhibits 10,11 body, selenium exerts function primarily as selenocysteine, cytoprotective, anti-inammatory, and antioxidant effects. oen referred to as the 21st amino acid, which is incorporated Similarly, the glutathione-mediated reduction of selenite 2À into and gives rise to the oen unique reactivity of (SeO3 ) to elemental selenium is thought to proceed through selenoproteins.7 a selenodiglutathione (GS-Se-SG) intermediate en-route to Twenty-ve selenoproteins have been identied in humans a selenopersulde (GS-SeH), which subsequently either 0 and fall into several main categories. These categories include decomposes to GSH and Se or is converted to H2Se through 12,13 glutathione peroxidases (Gpx), which scavenge harmful both enzymatic and non-enzymatic pathways. More recently, peroxide species, thioredoxin reductases (TrxR), which regulate the hydrolysis of phthalic selenoanhydride was used to generate thiol-disulde redox homeostasis, iodothyronine deiodinases reactive selenium species (RSeS) to examine the differential (DIOs), which regulate thyroid hormone equilibria, and synergies of these compounds with H2S and GSH in radical 14 specialized selenoproteins that exhibit alternative functions, scavenging. In this investigation, H2Se release was proposed such as protein folding and selenium transport.3 Selenium- during hydrolysis but was not observed directly in the experiments. Interest in developing chemical tools for investigating H2Se Department of Chemistry and Biochemistry, Materials Science Institute, Institute of and related RSeS has grown in the last few years, with new Molecular Biology, University of Oregon, Eugene, OR 97403, USA. E-mail: pluth@ À uoregon.edu investigations into the molecular recognition of HSe in 15 † Electronic supplementary information (ESI) available: Synthetic details, NMR synthetic host-guest systems and with the advent of rst- 16,17 spectra, HPLC experiments, crystallographic information. CCDC 1953366. For generation uorescent probes for H2Se detection. In part, ESI and crystallographic data in CIF or other electronic format see DOI: these investigations are motivated by potential roles of 10.1039/c9sc04616j This journal is © The Royal Society of Chemistry 2019 Chem. Sci.,2019,10,10723–10727 | 10723 View Article Online Chemical Science Edge Article Creative Commons Attribution-NonCommercial 3.0 Unported Licence. Fig. 1 Schematic representation of metabolic pathways of dietary selenium compounds including various reactive selenium species. Selenate 2À 2À À (SeO4 ), selenite (SeO3 ), selenophosphate (H2SePO3 ), selenocysteine (Sec), selenomethionine (SeMet), methylselenocysteine (MeSec), thioredoxin (Trx), thioredoxin reductase (TrxR), glutaredoxin (Grx), glutathione reductase (GR), glutathione (GSH), diglutathione (GSSG), sele- nodiglutathione (GSSeSG), glutathioselenol (GSSeH), selenophosphate synthetase (SPS), selenocysteine lyase (SCLY), S-adenosylmethionine (SAM), transsulfuration pathway (TSP), elemental selenium (Se0), and reactive oxygen species (ROS). 23 This article is licensed under a selenides in treating conditions ranging from arsenic species and gasotransmitter. Substantial efforts have focused poisoning, in which toxic arsenic species can react with H2Se to on the development of small-molecule H2S donors for delivery 18,19 24–26 form readily-excreted products, to cancer, in which H2Se can to biological environments. Although the structure and induce oxidative stress under normoxic conditions or reductive complexity of such systems have evolved signicantly, an early Open Access Article. Published on 11 October 2019. Downloaded 9/28/2021 8:08:42 AM. stress under hypoxic conditions in HepG2 cells, resulting in and broadly-used example of such donors is the hydrolysis- HMGB1 protein damage and ultimately apoptosis.20 At physio- activated donor GYY4137, which relies on the hydrolytic À ] 27 logical pH, almost all H2Se exists as HSe due to the acidity of cleavage of P S bonds to generate H2S. GYY4137 has been ¼ the diprotic form of H2Se (pKa 3.9). The high redox activity used in >200 publications to date (Web of Science) and exhibits À and high nucleophilicity of H2Se/HSe , when coupled with the À relatively low biological selenium content (0.2 mg kg 1 in humans), make investigations into the biological roles of H2Se difficult.21,22 Building from these past results and increased interest in biorelevant small RSeS, we viewed that well- characterized, synthetic small molecules that release H2Se directly and under controlled conditions would provide a much- needed chemical tool for expanding research related to the chemical biology of selenium. Here we report the development and characterization of a hydrolysis-based small-molecule H2Se donor and provide insights into the reaction mechanism and methods for direct H2Se trapping. Results and discussion Drawing parallels to biological organosulfur chemistry, the last Fig. 2 (a) Synthesis of TDN1042 from Woollins' reagent. (b) ORTEP een years have witnessed a surge in research related to diagram (50% ellipsoids) of TDN1042. Hydrogen atoms, except those À hydrogen sulde (H2S/HS ) as an important reactive sulfur on the morpholinium nitrogen, are omitted for clarity. 10724 | Chem. Sci.,2019,10,10723–10727 This journal is © The Royal Society of Chemistry 2019 View Article Online Edge Article Chemical Science anti-inammatory, vasorelaxant, and anti-cancer as well as measured the rate of hydrolysis of TDN1042 (10 mM) in citrate other effects in different biological models27–29 with diverse buffer (50 mM) ranging from pH 3.0 to pH 6.0 in ame-sealed applications ranging from medicinal to agricultural science.30,31 NMR tubes at ambient temperature (Fig. 3b). The resulting Motivated by the broad utility of this approach to access H2S hydrolysis data (Fig. 3c) revealed an increase in rate at more donor motifs, we sought to use similar chemistry to generate acidic pH values, which is consistent with the expected hydro- well-dened H2Se donors that are activated by P]Se bond lysis mechanism. A similar pH dependence was observed for hydrolysis. To prepare such a donor, we treated Woollins' GYY4137 in a previous report,27 although the experimental reagent with an excess of morpholine, drawing parallels to the conditions and methods used to monitor rates and product synthesis of GYY4137, to generate TDN1042 in moderate yield conversions are too dissimilar to those used here for TDN1042 (Fig. 2a). The resultant product was characterized by 1H, 13C to make direct quantitative comparisons. This similarity in pH {1H}, 31P, and 77Se NMR spectroscopy (Fig. S1–S4†). Single