[biochemical] transport of hydrogen sulfide (H2S) activated by cysteine via cage like carbonyl sulfur (COS) donor

Since endogenous H 2S was proved to be a neuroactive substance in 1996, H 2S has been added to the family of gas signaling molecules H 2S is mainly produced by four enzymes: cystathionine β - synthetase (CBS) and cystathionine γ - lyase (CSE) produce endogenous H 2S, and cysteine (Cys) and homocysteine (Hcy) are transformed into H 2S H 2 S is involved in many pathophysiological processes and plays an important role in anti-inflammatory, reducing oxidative stress and vasodilation Therefore, the development of H 2S releaser (H 2S donor) has been one of the hot research fields Although sulfurized salts such as sodium hydrosulfide (NaHS) and sodium sulfide (Na2S) have been widely used in this field, they can spontaneously release H2S, which leads to the rapid oxidation of H2S Therefore, sulfurized salts are far from ideal donors of H2S Therefore, it is a key tool for H 2S research to find stable and controllable H 2S donor According to this demand, the Michael pluth group of the University of Oregon reported the study of cage like carbonyl sulfur (COS) as a new type of H 2S donor molecule Compared with other known H 2S donor molecules, cos based donors release cos after being excited, and COS can be rapidly converted into H 2S under the catalysis of carbonic anhydrase (CA) In addition, the team also found that caged thiocarbamates and thiocarbonates are also promising as cos donors An important advantage of this strategy is that it can design cos release scaffold under certain conditions to realize the transport of H 2S Initially, the team and other scientists applied this strategy to other donor molecules, and excited them through different trigger mechanisms, such as ROS, esterase, nucleophile, click chemistry and light (Fig 1) (source: Chemical Communications) nucleophiles play an important role in biological systems, among which thiols such as Cys and reduced glutathione (GSH) have attracted more attention due to cell abundance and potential reactivity Cys and GSH have been widely used to trigger bioactive molecules and prodrugs, and then release cage compounds, including sulfur dioxide (SO2), nitroacyl (HNO) and anticancer drugs It is important that thiol activation has been applied to the development of H 2S donors and several unstable H 2S donors, and has shown the potential of protection in the clinical trials of animal models Inspired by these studies, Michael pluth team first reported the CoS / H 2S donor excited by Cys, and used acrylate with Cys activity to functionalize thiocarbamate The author found that these thiocarbamates can not only expand the current cos based H 2S donor molecules, but also serve as a powerful tool for H 2S research 10.1039/c8cc02428f ）。 Cys can react with acrylate to produce substituted 1,4-thioaza compounds Firstly, Cys attacked acrylate to produce thioether, and then through intramolecular cyclization to produce 1,4-thioaza compounds Strongin et al Used this cyclization strategy to design a fluorescence probe based on Acrylate for Cys detection Recently, berreau group has developed a series of CO donor molecules with similar response Based on the current research situation, the author uses Cys acrylate reaction as a trigger mechanism to obtain a new CoS / H 2S donor, which is activated by cycloaddition / cyclization mediated by Cys The resulting phenol intermediate undergoes a 1,6-elimination reaction to release cos, which can be quickly converted into H 2S (scheme1) in the presence of Ca enzyme (source: Chemical Communications) in order to verify the corresponding assumption, the author prepared o-alkyltyrosine sensitive thiocarbamate molecule (oa-cystcm-1) Once activated, oa-cystcm-1 will release cos, which can be rapidly hydrogenated to H 2S in the presence of Ca enzyme In addition, the corresponding control molecules, oa-cyscm-1 and oa-tcm-1, were designed Oa-cyscm-1 is obtained by the reaction of 4-hydroxymethylphenylacrylate with phenyl isocyanate The authors predict that oa-cyscm-1 can release CO2 instead of COS after the same Cys activation On the other hand, oa-tcm-1 lacks acrylate trigger due to the retention of thiocarbamate scaffold, so it will not react with Cys and it is impossible to release cos (scheme2) in other ways (source: Chemical Communications) the authors used methylene blue (MB) analysis to monitor the release of H 2S from oa-cystcm-1 (50mm) in the presence of Cys (0-500mm) Cys was dissolved in PBS buffer containing ca In the absence of Cys, oa-cystcm-1 can exist stably in the aqueous phase and cannot spontaneously release CoS / H2S; however, after adding Cys, it will release CoS / H2S, and this release is dose-dependent (Fig 2) The experimental results show that oa-cystcm-1 can be excited by Cys, and the cos can be transformed into H2S rapidly in the presence of ca In order to further explain that the release of H 2S was initiated by Cys, the author used N-ethylmaleimide (NEM), a Cys scavenger, to pretreat Cys, and then added oa-cystcm-1 Compared with the conventional excitation conditions, NEM pretreatment can significantly eliminate the release of H2S from oa-cystcm-1, which further indicates that the activation of receptor requires the presence of Cys; only in the presence of Ca, after Ca pretreatment with acetazolamide (AAA, a CA inhibitor), no release of H2S was observed, which indicates that the release of H2S from oa-cystcm-1 is conducted in a cos dependent manner (Fig 3) (source: Chemical Communications) in addition to Cys, the author also tested the donor activity of other biological nucleophiles As predicted by the authors, when these molecules trigger oa-cystcm-1, there is no CoS / H2 S release, which is due to the lack of addition / cyclization excitation sequence The results showed that in the presence of both Cys and GSH, the release of H2S decreased with the increase of GSH concentration, which indicated that GSH could induce donor consumption Although the effect of GSH in aqueous solution is not obvious, it should also be considered when applying donor to biological system Oa-cystcm-1 was cultured with pig liver esterase (PLE) to determine whether the esterase can release CoS / H 2S Although the authors observed the release of H 2S, its efficiency was significantly lower than that triggered by Cys As the author expected, no production of H 2S was observed after the control compounds oa-cyscm-1 and oa-tcm-1 were treated with Cys in the presence of Ca, which indicated that the aryl acrylate trigger and thiocarbamate scaffold were crucial for the release of COS According to the current research, oa-cystcm-1 is highly sensitive to the activation of Cys, and then releases CoS / H2S, while it is inert to the activation of other biological molecules such as GSH, GSSG, Hcy, NAC, Ser and Lys Next, the release of CoS / H 2S from oa-cystcm-1 in the presence of Cys was studied at the cell level The visual analysis was carried out by means of end.3 cells and sf7-am In the absence of oa-cystcm-1, negligible fluorescence of sf7-am was observed, indicating that a small amount of endogenous H 2S could be produced in the boend 3 cells; in contrast, the addition of oa-cystcm-1 resulted in a significant increase in the fluorescence of sf7-am, indicating that oa-cystcm-1 could be activated by Cys and release H 2S at the cell level (Fig 4) These results indicate that oa-cystcm-1 is an effective CoS / H 2S donor and has potential application value in the treatment and research tools related to H 2S (source: Chemical Communications) in a word, the author reported the preparation of oa-cystcm-1, a CoS / H 2S donor triggered by Cys, and its release of H 2S The results showed that oa-cystcm-1 was stable in aqueous medium, and COS / H 2S was not released until it was activated by Cys Importantly, the transport of H 2S through oa-cystcm-1 can be observed at the cell level, which indicates that oa-cystcm-1 can also be an effective CoS / H 2S donor in complex biological systems The author's current research makes oa-cystcm-1 become a new member of the H2S donor family based on cos, and has potential application value in the fields of chemistry and biology related to H2S and COS.