Unexpected diselenide metathesis in selenocysteine-substituted biologically active peptides.

Substitution of disulfide bonds with a diselenide bonds in peptides and proteins is an often-used strategy to increase the stability of naturally occurring peptides and proteins. In this paper, diselenide metathesis between model diselenide dimer peptides, as well as that in diselenide(s)-substituted biologically active peptides, were analyzed. Surprisingly, depending on the tertiary structure of the peptides, we observed that the metathesis reaction occurs under physiological conditions even in the absence of reducing agents, light and heating.

Effect of exogenous selenium on physicochemical, structural, functional, thermal, and gel rheological properties of mung bean (Vigna radiate L.) protein.

Selenium (Se) biofortification during the growth process of mung bean is an effective method to improve the Se content and quality. However, the effect of Se biofortification on the physicochemical properties of mung bean protein is unclear. The objective of this study was to clarify the changes in the composition, Se forms, particle structure, functional properties, thermal stability, and gel properties of mung bean protein at four Se application levels. The results showed that the Se content of mung bean protein increased in a dose-dependent manner, with 7.96-fold (P1) and 8.52-fold (P2) enhancement at the highest concentration. Exogenous Se application promotes the conversion of inorganic Se to organic Se. Among them, selenomethionine (SeMet) and methyl selenocysteine (MeSeCys) replaced Met and Cys through the S metabolic pathway and became the dominant organic Se forms in Se-enriched mung bean protein, accounting for more than 80 % of the total Se content. Exogenous Se at 30 g/hm2 significantly up-regulated protein content and promoted the synthesis of sulfur-containing protein components and hydrophobic amino acids in the presence of increased levels of SeMet and MeSeCys. Meanwhile, Cys and Met substitution altered the sulfhydryl groups (SH), ß-sheets, and ß-turns of protein. The particle size and microstructural characteristics depend on the protein itself and were not affected by exogenous Se. The Se-induced increase in the content of hydrophobic amino acids and ß-sheets synergistically increases the thermal stability of the protein. Moderate Se application altered the functional properties of mung bean protein, which was mainly reflected in the significant increase in oil holding capacity (OHC) and foaming capacity (FC). In addition, the increase in SH and ß-sheets induced by exogenous Se could alter the protein intermolecular network, contributing to the increase in storage modulus (G') and loss modulus (G″), which resulted in the formation of more highly elastic gels. This study further promotes the application of mung bean protein in the field of food processing and provides a theoretical basis for the extensive development of Se-enriched mung bean protein.

Enzymatic strategies for selenium incorporation into biological molecules.

The trace element selenium (Se) is essential to the physiology of most organisms on the planet. The most well documented of Se's biological forms are selenoproteins, where selenocysteine often serves as the catalytic center for crucial redox processes. Se is also found in several other classes of biological molecules, including nucleic acids, sugars, and modified amino acids, although its role in the function of these metabolites is less understood. Despite its prevalence, only a small number of Se-specific biosynthetic pathways have been discovered. Around half of these were first characterized in the past three years, suggesting that the selenometabolome may be more diverse than previously appreciated. Here, we review the recent advances in our understanding of this intriguing biochemical space, and discuss prospects for future discovery efforts.

Asgard archaeal selenoproteome reveals a roadmap for the archaea-to-eukaryote transition of selenocysteine incorporation machinery.

Selenocysteine (Sec) is encoded by the UGA codon that normally functions as a stop signal and is specifically incorporated into selenoproteins via a unique recoding mechanism. The translational recoding of UGA as Sec is directed by an unusual RNA structure, the SECIS element. Although archaea and eukaryotes adopt similar Sec encoding machinery, the SECIS elements have no similarities to each other with regard to sequence and structure. We analyzed >400 Asgard archaeal genomes to examine the occurrence of both Sec encoding system and selenoproteins in this archaeal superphylum, the closest prokaryotic relatives of eukaryotes. A comprehensive map of Sec utilization trait has been generated, providing the most detailed understanding of the use of this nonstandard amino acid in Asgard archaea so far. By characterizing the selenoproteomes of all organisms, several selenoprotein-rich phyla and species were identified. Most Asgard archaeal selenoprotein genes possess eukaryotic SECIS-like structures with varying degrees of diversity. Moreover, euryarchaeal SECIS elements might originate from Asgard archaeal SECIS elements via lateral gene transfer, indicating a complex and dynamic scenario of the evolution of SECIS element within archaea. Finally, a roadmap for the transition of eukaryotic SECIS elements from archaea was proposed, and selenophosphate synthetase may serve as a potential intermediate for the generation of ancestral eukaryotic SECIS element. Our results offer new insights into a deeper understanding of the evolution of Sec insertion machinery.

PRDX6 augments selenium utilization to limit iron toxicity and ferroptosis.

Ferroptosis is a form of regulated cell death induced by iron-dependent accumulation of lipid hydroperoxides. Selenoprotein glutathione peroxidase 4 (GPX4) suppresses ferroptosis by detoxifying lipid hydroperoxides via a catalytic selenocysteine (Sec) residue. Sec, the genetically encoded 21st amino acid, is biosynthesized from a reactive selenium donor on its cognate tRNA[Ser]Sec. It is thought that intracellular selenium must be delivered 'safely' and 'efficiently' by a carrier protein owing to its high reactivity and very low concentrations. Here, we identified peroxiredoxin 6 (PRDX6) as a novel selenoprotein synthesis factor. Loss of PRDX6 decreases the expression of selenoproteins and induces ferroptosis via a reduction in GPX4. Mechanistically, PRDX6 increases the efficiency of intracellular selenium utilization by transferring selenium between proteins within the selenocysteyl-tRNA[Ser]Sec synthesis machinery, leading to efficient synthesis of selenocysteyl-tRNA[Ser]Sec. These findings highlight previously unidentified selenium metabolic systems and provide new insights into ferroptosis.

Se-methylselenocysteine inhibits the progression of non-small cell lung cancer via ROS-mediated NF-κB signaling pathway.

Se-methylselenocysteine (MSC) is recognized for its potential in cancer prevention, yet the specific effects and underlying processes it initiates within non-small cell lung cancer (NSCLC) remain to be fully delineated. Employing a comprehensive array of assays, including CCK-8, colony formation, flow cytometry, MitoSOX Red staining, wound healing, transwell, and TUNEL staining, we evaluated MSC's effects on A549 and 95D cell lines. Our investigation extended to the ROS-mediated NF-κB signaling pathway, utilizing Western blot analysis, P65 overexpression, and the application of IκB-α inhibitor (BAY11-7082) or N-acetyl-cysteine (NAC) to elucidate MSC's mechanism of action. In vivo studies involving subcutaneous xenografts in mice further confirmed MSC's inhibitory effect on tumor growth. Our findings indicated that MSC inhibited the proliferation of A549 and 95D cells, arresting cell cycle G0/G1 phase and reducing migration and invasion, while also inducing apoptosis and increasing intracellular ROS levels. This was accompanied by modulation of key proteins, including the upregulation of p21, p53, E-cadherin, Bax, cleaved caspase-3, cleaved-PARP, and downregulation of CDK4, SOD2, GPX-1. MSC was found to inhibit the NF-κB pathway, as evidenced by decreased levels of P-P65 and P-IκBα. Notably, overexpression of P65 and modulation of ROS levels with NAC could attenuate MSC's effects on cellular proliferation and metastasis. Moreover, MSC significantly curtailed tumor growth in vivo and disrupted the NF-κB signaling pathway. In conclusion, our research demonstrates that MSC exhibits anticancer effects against NSCLC by modulating the ROS/NF-κB signaling pathway, suggesting its potential as a therapeutic agent in NSCLC treatment.

Glutathione peroxidase (GPX1) - Selenocysteine metabolism preserves the follicular fluid's (FF) redox homeostasis via IGF-1- NMD cascade in follicular ovarian cysts (FOCs).

Follicular ovarian cysts (FOCs) are characterized by follicles in the ovaries that are >20 mm in diameter and persist for >10 days without the corpus luteum, leading to anovulation, dysregulation of folliculogenesis and subfertility in humans and livestock species. Despite their clinical significance, the precise impact of FOCs on oocyte reserve, maturation, and quality still needs to be explored. While FOCs are observed in both human and livestock populations, they are notably prevalent in livestock species. Consequently, livestock species serve as valuable models for investigating the molecular intricacies of FOCs. Thus, in this study, using goat FOCs, we performed integrated proteomic, metabolomic and functional analyses to demonstrate that oocyte maturation is hampered due to increased reactive oxygen species (ROS) in FOCs follicular fluid (FF) via downregulation of glutathione peroxidase (GPX1), a critical antioxidant seleno enzyme required to negate oxidative stress. Notably, GPX1 reduction was positively correlated with the FF's decline of free selenium and selenocysteine metabolic enzymes, O-phosphoryl-tRNA (Sec) selenium transferase (SEPSECS) and selenocysteine lyase (SCLY) levels. Adding GPX1, selenocysteine, or selenium to the culture media rescued the oocyte maturation abnormalities caused by FOCs FF by down-regulating the ROS. Additionally, we demonstrate that substituting GPX1 regulator, Insulin-like growth factor-I (IGF-1) in the in vitro maturation media improved the oocyte maturation in the cystic FF by down-regulating the ROS activity via suppressing Non-sense-mediated decay (NMD) of GPX1. In contrast, inhibition of IGF-1R and the target of rapamycin complex 1 (mTORC1) hampered the oocyte maturation via NMD up-regulation. These findings imply that the GPX1 regulation via selenocysteine metabolism and the IGF-1-mediated NMD may be critical for the redox homeostasis of FF. We propose that GPX1 enhancers hold promise as therapeutics for enhancing the competence of FOCs oocytes. However, further in vivo studies are necessary to validate these findings observed in vitro.

Replacing a Cysteine Ligand by Selenocysteine in a [NiFe]-Hydrogenase Unlocks Hydrogen Production Activity and Addresses the Role of Concerted Proton-Coupled Electron Transfer in Electrocatalytic Reversibility.

Hydrogenases catalyze hydrogen/proton interconversion that is normally electrochemically reversible (having minimal overpotential requirement), a special property otherwise almost exclusive to platinum metals. The mechanism of [NiFe]-hydrogenases includes a long-range proton-coupled electron-transfer process involving a specific Ni-coordinated cysteine and the carboxylate of a nearby glutamate. A variant in which this cysteine has been exchanged for selenocysteine displays two distinct changes in electrocatalytic properties, as determined by protein film voltammetry. First, proton reduction, even in the presence of H2 (a strong product inhibitor), is greatly enhanced relative to H2 oxidation: this result parallels a characteristic of natural [NiFeSe]-hydrogenases which are superior H2 production catalysts. Second, an inflection (an S-shaped "twist" in the trace) appears around the formal potential, the small overpotentials introduced in each direction (oxidation and reduction) signaling a departure from electrocatalytic reversibility. Concerted proton-electron transfer offers a lower energy pathway compared to stepwise transfers. Given the much lower proton affinity of Se compared to that of S, the inflection provides compelling evidence that concerted proton-electron transfer is important in determining why [NiFe]-hydrogenases are reversible electrocatalysts.

[Experimental study on promotion of peripheral nerve regeneration by selenium-methylselenocysteine].

Objective: To investigate the feasibility of selenium-methylselenocysteine (SMC) to promote peripheral nerve regeneration and its mechanism of action. Methods: Rat Schwann cells RSC96 cells were randomly divided into 5 groups, which were group A (without any treatment, control group), group B (adding 100 µmol/L H 2O 2), group C (adding 100 µmol/L H 2O 2+100 µmol/L SMC), group D (adding 100 µmol/L H 2O 2+200 µmol/L SMC), group E (adding 100 µmol/L H 2O 2+400 µmol/L SMC); the effect of SMC on cell proliferation was detected by MTT method, and the level of oxidative stress was detected by immunofluorescence for free radicals [reactive oxygen species (ROS)] after determining the appropriate dose group. Thirty-six 4-week-old male Sprague Dawley rats were randomly divided into 3 groups, namely, the sham operation group (Sham group), the sciatic nerve injury group (PNI group), and the SMC treatment group (SMC group), with 12 rats in each group; the rats in the PNI group were fed with food and water normally after modelling operation, and the rats in the SMC group were added 0.75 mg/kg SMC to the drinking water every day. At 4 weeks after operation, the sciatic nerves of rats in each group were sampled for neuroelectrophysiological detection of highest potential of compound muscle action potential (CMAP). The levels of inflammatory factors [interleukin 17 (IL-17), IL-6, IL-10 and oxidative stress factors catalase (CAT), superoxide dismutase (SOD), and malondialdehyde (MDA)] were detected by ELISA assay. The luxol fast blue (LFB) staining was used to observe the myelin density, fluorescence intensity of glial fibrillary acidic protein (GFAP) and myelin basic protein (MBP) was observed by immunofluorescence staining, and myelin morphology was observed by transmission electron microscopy with measurement of axon diameter. Western blot was used to detect the protein expressions of p38 mitogen-activated protein kinases (p38MAPK), phosphorylated p38MAPK (p-p38MAPK), heme oxygenase 1 (HO-1), and nuclear factor erythroid 2-related factor 2 (Nrf2). Results: MTT assay showed that the addition of SMC significantly promoted the proliferation of RSC96 cells, and the low concentration could achieve an effective effect, so the treatment method of group C was selected for the subsequent experiments; ROS immunofluorescence test showed that group B showed a significant increase in the intensity of ROS fluorescence compared with that of group A, and group C showed a significant decrease in the intensity of ROS fluorescence compared with that of group B ( P<0.05). Neuroelectrophysiological tests showed that the highest potential of CMAP in SMC group was significantly higher than that in PNI and Sham groups ( P<0.05). ELISA assay showed that the levels of IL-6, IL-17, and MDA in PNI group were significantly higher than those in Sham group, and the levels of IL-10, SOD, and CAT were significantly lower; the levels of IL-6, IL-17, and MDA in SMC group were significantly lower than those in PNI group, and the levels of IL-10, SOD, and CAT were significantly higher ( P<0.05). LFB staining and transmission electron microscopy showed that the myelin density and the diameter of axons in the SMC group were significantly higher than those of the PNI group and the Sham group ( P<0.05). Immunofluorescence staining showed that the fluorescence intensity of GFAP and MBP in the SMC group were significantly stronger than those in the PNI group and Sham group ( P<0.05). Western blot showed that the relative expressions of Nrf2 and HO-1 proteins in the SMC group were significantly higher than those in the PNI group and Sham group, and the ratio of p-p38MAPK/p38MAPK proteins was significantly higher in the PNI group than that in the SMC group and Sham group ( P<0.05). Conclusion: SMC may inhibit oxidative stress and inflammation after nerve injury by up-regulating the Nrf2/HO-1 pathway, and then inhibit the phosphorylation of p38MAPK pathway to promote the proliferation of Schwann cells, which ultimately promotes the formation of myelin sheaths and accelerates the regeneration of peripheral nerves.

The Selenoproteome as a Dynamic Response Mechanism to Oxidative Stress in Hydrogenotrophic Methanogenic Communities.

Methanogenesis is a critical process in the carbon cycle that is applied industrially in anaerobic digestion and biogas production. While naturally occurring in diverse environments, methanogenesis requires anaerobic and reduced conditions, although varying degrees of oxygen tolerance have been described. Microaeration is suggested as the next step to increase methane production and improve hydrolysis in digestion processes; therefore, a deeper understanding of the methanogenic response to oxygen stress is needed. To explore the drivers of oxygen tolerance in methanogenesis, two parallel enrichments were performed under the addition of H2/CO2 in an environment without reducing agents and in a redox-buffered environment by adding redox mediator 9,10-anthraquinone-2,7-disulfonate disodium. The cellular response to oxidative conditions is mapped using proteomic analysis. The resulting community showed remarkable tolerance to high-redox environments and was unperturbed in its methane production. Next to the expression of pathways to mitigate reactive oxygen species, the higher redox potential environment showed an increased presence of selenocysteine and selenium-associated pathways. By including sulfur-to-selenium mass shifts in a proteomic database search, we provide the first evidence of the dynamic and large-scale incorporation of selenocysteine as a response to oxidative stress in hydrogenotrophic methanogenesis and the presence of a dynamic selenoproteome.

Ancient Loss of Catalytic Selenocysteine Spurred Convergent Adaptation in a Mammalian Oxidoreductase.

Selenocysteine, the 21st amino acid specified by the genetic code, is a rare selenium-containing residue found in the catalytic site of selenoprotein oxidoreductases. Selenocysteine is analogous to the common cysteine amino acid, but its selenium atom offers physical-chemical properties not provided by the corresponding sulfur atom in cysteine. Catalytic sites with selenocysteine in selenoproteins of vertebrates are under strong purifying selection, but one enzyme, glutathione peroxidase 6 (GPX6), independently exchanged selenocysteine for cysteine <100 million years ago in several mammalian lineages. We reconstructed and assayed these ancient enzymes before and after selenocysteine was lost and up to today and found them to have lost their classic ability to reduce hydroperoxides using glutathione. This loss of function, however, was accompanied by additional amino acid changes in the catalytic domain, with protein sites concertedly changing under positive selection across distant lineages abandoning selenocysteine in glutathione peroxidase 6. This demonstrates a narrow evolutionary range in maintaining fitness when sulfur in cysteine impairs the catalytic activity of this protein, with pleiotropy and epistasis likely driving the observed convergent evolution. We propose that the mutations shared across distinct lineages may trigger enzymatic properties beyond those in classic glutathione peroxidases, rather than simply recovering catalytic rate. These findings are an unusual example of adaptive convergence across mammalian selenoproteins, with the evolutionary signatures possibly representing the evolution of novel oxidoreductase functions.

Se-methylselenocysteine ameliorates mitochondrial function by targeting both mitophagy and autophagy in the mouse model of Alzheimer's disease.

Background: Alzheimer's disease (AD) exerts tremendous pressure on families and society due to its unknown etiology and lack of effective treatment options. Our previous study had shown that Se-methylselenocysteine (SMC) improved the cognition and synaptic plasticity of triple-transgenic AD (3 × Tg-AD) mice and alleviated the related pathological indicators. We are dedicated to investigating the therapeutic effects and molecular mechanisms of SMC on mitochondrial function in 3 × Tg-AD mice. Methods: Transmission electron microscopy (TEM), western blotting (WB), mitochondrial membrane potential (ΔΨm), mitochondrial swelling test, and mitochondrial oxygen consumption test were used to evaluate the mitochondrial morphology and function. Mitophagy flux and autophagy flux were assessed with immunofluorescence, TEM and WB. The Morris water maze test was applied to detect the behavioral ability of mice. Results: The destroyed mitochondrial morphology and function were repaired by SMC through ameliorating mitochondrial energy metabolism, mitochondrial biogenesis and mitochondrial fusion/fission balance in 3 × Tg-AD mice. In addition, SMC ameliorated mitochondria by activating mitophagy flux via the BNIP3/NIX pathway and triggering autophagy flux by suppressing the Ras/Raf/MEK/ERK/mTOR pathway. SMC remarkably increased the cognitive ability of AD mice. Conclusions: This research indicated that SMC might exert its therapeutic effect by protecting mitochondria in 3 × Tg-AD mice.

Mechanism of Action of Antitumor Au(I) N-Heterocyclic Carbene Complexes: A Computational Insight on the Targeting of TrxR Selenocysteine.

The targeting of human thioredoxin reductase is widely recognized to be crucially involved in the anticancer properties of several metallodrugs, including Au(I) complexes. In this study, the mechanism of reaction between a set of five N-heterocyclic carbene Au(I) complexes and models of the active Sec residue in human thioredoxin reductase was investigated by means of density functional theory approaches. The study was specifically addressed to the kinetics and thermodynamics of the tiled process by aiming at elucidating and explaining the differential inhibitory potency in this set of analogous Au(I) bis-carbene complexes. While the calculated free energy profile showed a substantially similar reactivity, we found that the binding of these Au(I) bis-carbene at the active CysSec dyad in the TrxR enzyme could be subjected to steric and orientational restraints, underlining both the approach of the bis-carbene scaffold and the attack of the selenol group at the metal center. A new and detailed mechanistic insight to the anticancer activity of these Au(I) organometallic complexes was thus provided by consolidating the TrxR targeting paradigm.

Engineered mRNA-ribosome fusions for facile biosynthesis of selenoproteins.

Ribosomes are often used in synthetic biology as a tool to produce desired proteins with enhanced properties or entirely new functions. However, repurposing ribosomes for producing designer proteins is challenging due to the limited number of engineering solutions available to alter the natural activity of these enzymes. In this study, we advance ribosome engineering by describing a novel strategy based on functional fusions of ribosomal RNA (rRNA) with messenger RNA (mRNA). Specifically, we create an mRNA-ribosome fusion called RiboU, where the 16S rRNA is covalently attached to selenocysteine insertion sequence (SECIS), a regulatory RNA element found in mRNAs encoding selenoproteins. When SECIS sequences are present in natural mRNAs, they instruct ribosomes to decode UGA codons as selenocysteine (Sec, U) codons instead of interpreting them as stop codons. This enables ribosomes to insert Sec into the growing polypeptide chain at the appropriate site. Our work demonstrates that the SECIS sequence maintains its functionality even when inserted into the ribosome structure. As a result, the engineered ribosomes RiboU interpret UAG codons as Sec codons, allowing easy and site-specific insertion of Sec in a protein of interest with no further modification to the natural machinery of protein synthesis. To validate this approach, we use RiboU ribosomes to produce three functional target selenoproteins in Escherichia coli by site-specifically inserting Sec into the proteins' active sites. Overall, our work demonstrates the feasibility of creating functional mRNA-rRNA fusions as a strategy for ribosome engineering, providing a novel tool for producing Sec-containing proteins in live bacterial cells.

Gold-Promoted Biocompatible Selenium Arylation of Small Molecules, Peptides and Proteins.

A low pKa (5.2), high polarizable volume (3.8 Å), and proneness to oxidation under ambient conditions make selenocysteine (Sec, U) a unique, natural reactive handle present in most organisms across all domains of life. Sec modification still has untapped potential for site-selective protein modification and probing. Herein we demonstrate the use of a cyclometalated gold(III) compound, [Au(bnpy)Cl2 ], in the arylation of diselenides of biological significance, with a scope covering small molecule models, peptides, and proteins using a combination of multinuclear NMR (including 77 Se NMR), and LC-MS. Diphenyl diselenide (Ph-Se)2 and selenocystine, (Sec)2 , were used for reaction optimization. This approach allowed us to demonstrate that an excess of diselenide (Au/Se-Se) and an increasing water percentage in the reaction media enhance both the conversion and kinetics of the C-Se coupling reaction, a combination that makes the reaction biocompatible. The C-Se coupling reaction was also shown to happen for the diselenide analogue of the cyclic peptide vasopressin ((Se-Se)-AVP), and the Bos taurus glutathione peroxidase (GPx1) enzyme in ammonium acetate (2 mM, pH=7.0). The reaction mechanism, studied by DFT revealed a redox-based mechanism where the C-Se coupling is enabled by the reductive elimination of the cyclometalated Au(III) species into Au(I).

A terbium(III) complex-based time-resolved luminescent probe for selenocysteine as an inhibitor of selenoproteins.

A terbium(III) complex-based time-resolved luminescence probe for selenocysteine can inhibit selenoprotein activity via a selenolate-triggered cleavage reaction of sulfonamide bonds in living cells.

Guiding bar motif of thioredoxin reductase 1 modulates enzymatic activity and inhibitor binding by communicating with the co-factor FAD and regulating the flexible C-terminal redox motif.

Thioredoxin reductase (TXNRD) is a selenoprotein that plays a crucial role in cellular antioxidant defense. Previously, a distinctive guiding bar motif was identified in TXNRD1, which influences the transfer of electrons. In this study, utilizing single amino acid substitution and Excitation-Emission Matrix (EEM) fluorescence spectrum analysis, we discovered that the guiding bar communicates with the FAD and modulates the electron flow of the enzyme. Differential Scanning Fluorimetry (DSF) analysis demonstrated that the aromatic amino acid in guiding bar is a stabilizer for TXNRD1. Kinetic analysis revealed that the guiding bar is vital for the disulfide reductase activity but hinders the selenocysteine-independent reduction activity of TXNRD1. Meanwhile, the guiding bar shields the selenocysteine residue of TXNRD1 from the attack of electrophilic reagents. We also found that the inhibition of TXNRD1 by caveolin-1 scaffolding domain (CSD) peptides and compound LCS3 did not bind to the guiding bar motif. In summary, the obtained results highlight new aspects of the guiding bar that restrict the flexibility of the C-terminal redox motif and govern the transition from antioxidant to pro-oxidant.

Maternal seafood consumption is associated with improved selenium status: Implications for child health.

Selenium (Se) is required for synthesis of selenocysteine (Sec), an amino acid expressed in the active sites of Se-dependent enzymes (selenoenzymes), including forms with essential functions in fetal development, brain activities, thyroid hormone metabolism, calcium regulation, and to prevent or reverse oxidative damage. Homeostatic mechanisms normally ensure the brain is preferentially supplied with Se to maintain selenoenzymes, but high methylmercury (CH3Hg) exposures irreversibly inhibit their activities and impair Sec synthesis. Due to Hg's high affinity for sulfur, CH3Hg initially binds with the cysteine (Cys) moieties of thiomolecules which are selenoenzyme substrates. These CH3Hg-Cys adducts enter selenoenzyme active sites and transfer CH3Hg to Sec, thus irreversibly inhibiting their activities. High CH3Hg exposures are uniquely able to induce a conditioned Se-deficiency that impairs synthesis of brain selenoenzymes. Since the fetal brain lacks Se reserves, it is far more vulnerable to CH3Hg exposures than adult brains. This prompted concerns that maternal exposures to CH3Hg present in seafood might impair child neurodevelopment. However, typical varieties of ocean fish contain far more Se than CH3Hg. Therefore, eating them should augment Se-status and thus prevent Hg-dependent loss of fetal selenoenzyme activities. To assess this hypothesis, umbilical cord blood and placental tissue samples were collected following delivery of a cohort of 100 babies born on Oahu, Hawaii. Dietary food frequency surveys of the mother's last month of pregnancy identified groups with no (0 g/wk), low (0-12 g/wk), or high (12 + g/wk) levels of ocean fish consumption. Maternal seafood consumption increased Hg contents in fetal tissues and resulted in ∼34% of cord blood samples exceeding the EPA Hg reference level of 5.8 ppb (0.029 µM). However, Se concentrations in these tissues were orders of magnitude higher and ocean fish consumption caused cord blood Se to increase ∼9.4 times faster than Hg. Therefore, this study supports the hypothesis that maternal consumption of typical varieties of ocean fish provides substantial amounts of Se that protect against Hg-dependent losses in Se bioavailability. Recognizing the pivotal nature of the Hg:Se relationship provides a consilient perspective of seafood benefits vs. risks and clarifies the reasons for the contrasting findings of certain early studies.

Se-(Methyl)-selenocysteine ameliorates blood-brain barrier disruption of focal cerebral ischemia mice via ferroptosis inhibition and tight junction upregulation in an Akt/GSK3ß-dependent manner.

BACKGROUND: Ischemic stroke still ranks as the most fatal disease worldwide. Blood-brain barrier (BBB) is a promising therapeutic target for protection. Brain microvascular endothelial cell is a core component of BBB, the barrier function maintenance of which can ameliorate ischemic injury and improve neurological deficit. Se-methyl L-selenocysteine (SeMC) has been shown to exert cardiovascular protection. However, the protection of SeMC against ischemic stroke remains to be elucidated. This research was designed to explore the protection of SeMC from the perspective of BBB protection. METHODS: To simulate cerebral ischemic injury, C57BL/6J mice were subjected to middle cerebral artery occlusion/reperfusion (MCAO/R), and bEnd.3 was exposed to oxygen-glucose deprivation/reoxygenation (OGD/R). After the intervention of SeMC, the barrier function and the expression of tight junction and ferroptosis-associated proteins were determined. For mechanism exploration, LY294002 (Akt inhibitor) was introduced both in vivo and in vitro. RESULTS: SeMC lessened the brain infarct volume and attenuated the leakage of BBB in mice. In vitro, SeMC improved cell viability and maintained the barrier function of bEnd.3 cells. The protection of SeMC was accompanied with ferroptosis inhibition and tight junction protein upregulation. Mechanism studies revealed that the effect of SeMC was reversed by LY294002, indicating that the protection of SeMC against ischemic stroke was mediated by the Akt signal pathway. CONCLUSION: These results suggested that SeMC exerted protection against ischemic stroke, which might be attributed to activating the Akt/GSK3ß signaling pathway and increasing the nuclear translocation of Nrf2 and ß-catenin, subsequently maintaining the integrity of BBB.

Se-Glargine: Chemical Synthesis of a Basal Insulin Analogue Stabilized by an Internal Diselenide Bridge.

Insulin has long provided a model for studies of protein folding and stability, enabling enhanced treatment of diabetes mellitus via analogue design. We describe the chemical synthesis of a basal insulin analogue stabilized by substitution of an internal cystine (A6-A11) by a diselenide bridge. The studies focused on insulin glargine (formulated as Lantus® and Toujeo®; Sanofi). Prepared at pH 4 in the presence of zinc ions, glargine exhibits a shifted isoelectric point due to a basic B chain extension (ArgB31 -ArgB32 ). Subcutaneous injection leads to pH-dependent precipitation of a long-lived depot. Pairwise substitution of CysA6 and CysA11 by selenocysteine was effected by solid-phase peptide synthesis; the modified A chain also contained substitution of AsnA21 by Gly, circumventing acid-catalyzed deamidation. Although chain combination of native glargine yielded negligible product, in accordance with previous synthetic studies, the pairwise selenocysteine substitution partially rescued this reaction: substantial product was obtained through repeated combination, yielding a stabilized insulin analogue. This strategy thus exploited both (a) the unique redox properties of selenocysteine in protein folding and (b) favorable packing of an internal diselenide bridge in the native state, once achieved. Such rational optimization of protein folding and stability may be generalizable to diverse disulfide-stabilized proteins of therapeutic interest.