Selenium Decipher: Trapping of Native Selenomethionine-Containing Peptides in Selenium-Enriched Milk and Unveiling the Deterioration after Ultrahigh-Temperature Treatment.

Selenopeptide identification relies on databases to interpret the selenopeptide spectra. A common database search strategy is to set selenium as a variable modification instead of sulfur on peptides. However, this approach generally detects only a fraction of selenopeptides. An alternative approach, termed Selenium Decipher, is proposed in the present study. It involves identifying collision-induced dissociation-cleavable selenomethionine-containing peptides by iteratively matching the masses of seleno-amino acids in selenopeptide spectra. This approach uses variable-data-independent acquisition (vDIA) for peptide detection, providing a flexible and customizable window for secondary mass spectral fragmentation. The attention mechanism was used to capture global information on peptides and determine selenomethionine-containing peptide backbones. The core structure of selenium on selenomethionine-containing peptides generates a series of fragment ions, namely, C3H7Se+, C4H10NSe+, C5H7OSe+, C5H8NOSe+, and C7H11N2O2Se+, with known mass gaps during higher-energy collisional dissociation (HCD) fragmentation. De-selenium spectra are generated by removing selenium originating from selenium replacement and then reassigning the precursors to peptides. Selenium-enriched milk is obtained by feeding selenium-rich forage fed to cattle, which leads to the formation of native selenium through biotransformation. A novel antihypertensive selenopeptide Thr-Asp-Asp-Ile-SeMet-Cys-Val-Lys TDDI(Se)MCVK was identified from selenium-enriched milk. The selenopeptide (IC50 = 60.71 µM) is bound to four active residues of the angiotensin-converting enzyme (ACE) active pocket (Ala354, Tyr523, His353, and His513) and two active residues of zinc ligand (His387 and Glu411) and exerted a competitive inhibitory effect on the spatial blocking of active sites. The integration of vDIA and the iteratively matched seleno-amino acids was applied for Selenium Decipher, which provides high validity for selenomethionine-containing peptide identification.

Selenomethionine Inhibits NF-κB-mediated Inflammatory Responses of Bovine Mammary Epithelial Cells Caused by Klebsiella pneumoniae by Increasing Autophagic Flux.

Klebsiella pneumoniae (K. pneumoniae) is one of the major pathogens causing bovine clinical mastitis. Autophagy maintains cellular homeostasis and resists excessive inflammation in eukaryotic organisms. Selenomethionine (Se-Met) is commonly used as a source of selenium supplementation for dairy cows. This study aimed to investigate the effects of Se-Met on inflammatory responses mediated by nuclear factor-kappa B (NF-κB) through autophagy. We infected bovine mammary epithelial cell line (MAC-T) with K. pneumoniae and examined the expression of autophagy-related proteins and changes in autophagic vesicles, LC3 puncta, and autophagic flux at various intervals. The results showed that K. pneumoniae activated the early-stage autophagy of MAC-T cells. The levels of LC3-II, Beclin1, and ATG5, as well as the number of LC3 puncta and autophagic vesicles, increased after 2 h post-treatment. However, the late-stage autophagic flux was blocked. Furthermore, the effect of autophagy on NF-κB-mediated inflammation was investigated with different autophagy levels. The findings showed that enhanced autophagy inhibited the K. pneumoniae-induced inflammatory responses of MAC-T cells. The opposite results were found with the inhibition of autophagy. Finally, we examined the effect of Se-Met on NF-κB-mediated inflammation based on autophagy. The results indicated that Se-Met alleviated K. pneumoniae-induced autophagic flux blockage, inhibited NF-κB-mediated inflammation, and decreased the adhesion of K. pneumoniae to MAC-T cells. The inhibitory effect of Se-Met on NF-κB-mediated inflammation could be partially blocked by the autophagy inhibitor chloroquine (CQ). Overall, Se-Met attenuated K. pneumoniae-induced NF-κB-mediated inflammatory responses by enhancing autophagic flux.

SeMet alleviates LPS-induced eggshell gland necroptosis mediated inflammation by regulating the Keap1/Nrf2/HO-1 pathway.

Exposure to lipopolysaccharide (LPS) can lead to inflammation in a variety of tissues and organs. Selenium (Se) plays a crucial role in mitigating inflammatory damage. Compared with inorganic selenium, organic selenium, such as selenomethionine (SeMet), has the advantages of a higher absorption rate and lower toxicity in animals. This study examined the protective effects of SeMet on eggshell gland tissue damage caused by LPS. Hy-Line Brown laying hens were chosen as the experimental animals and were randomly assigned to four groups: control group (C), lipopolysaccharide group (LPS), SeMet group (Se), and SeMet + lipopolysaccharide group (Se + LPS). H&E staining and transmission electron microscope were performed to observe the pathological changes of eggshell glands, oxidative stress related indicators were measured using relevant kits, qRT‒PCR and western blotting were used to evaluate the mRNA and protein levels of the Nrf2 pathway, necroptosis, and inflammation related indicators. The results showed that LPS treatment increased the content of malondialdehyde (MDA), decreased the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), and decreased the content of glutathione (GSH). LPS increased the levels of Keap1, RIPK1, RIPK3, MLKL, TNF-α, COX-2, and NF-κB, while decreasing the levels of HO-1, NQO1, Nrf2, and Caspase-8. However, SeMet treatment effectively reversed the changes of the above indicators, indicating that SeMet alleviates eggshell gland cell necroptosis-mediated inflammation induced by LPS via regulating the Keap1/Nrf2/HO-1 pathway. This study elucidated the mechanism by which SeMet alleviates LPS-induced eggshell gland tissue damage in Hy-Line Brown laying hens and provided a new direction for expanding the application of SeMet in the feeding and production of laying hens.

Protective effect of selenomethionine on kidney injury induced by ochratoxin A in rabbits.

The purpose of this study was to investigate the protective effect and mechanism of selenomethionine (SeMet) on ochratoxin A (OTA)-induced nephrotoxicity in rabbits. In total, sixty Ira rabbits were randomly divided into 5 groups (the control group, OTA group, 0. 2 mg/kg SeMet + OTA group, 0. 4 mg/kg SeMet + OTA group, and 0. 6 mg/kg SeMet + OTA group). The rabbits were fed diets supplemented with different doses of SeMet for 21 days and given 0. 2 mg/kg OTA starting on day 15 for a week. The results showed that the SeMet supplementation could improve the changes in blood physiological indices and renal function decline caused by OTA poisoning, and alleviate pathological kidney injury in the rabbits. SeMet also increased the activities of total antioxidant capacity, superoxide dismutase, and glutathione peroxidase, and decreased the contents of malondialdehyde and reactive oxygen species and the expression of interleukin-1ß, interleukin-6, and tumor necrosis factor-α in the damaged kidneys of the rabbits. In addition, the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream gene heme oxygenase 1 (HO-1) was also inhibited after OTA poisoning, while SeMet activated the Nrf2 signaling pathway and enhanced the expression of Nrf2 and the downstream gene HO-1. In conclusion, SeMet protected against kidney injury caused by OTA in rabbits, and the mechanism may be the activation of the Nrf2 signaling pathway.

Dietary supplementation with selenomethionine enhances antioxidant capacity and selenoprotein gene expression in layer breeder roosters.

This study's objective was to investigate the effects of dietary Se (in the form of selenomethionine) on the antioxidant activity and selenoprotein gene expressions in layer breeder roosters. One hundred and eighty, 36-wk-old Jingfen layer breeder roosters were randomly allocated to one of 5 dietary treatments (0, 0.25, 0.5, 1, or 2 mg/kg Se) for 6 wk on a corn-soybean meal-based diet. Antioxidant parameters and selenoprotein gene expressions were assessed at the end of the experiment. The results showed that Se supplementation significantly increased the activity of T-SOD, CAT, GSH-Px, and superoxide anion scavenging ability in plasma (P ≤ 0.05), and activities of T-SOD, CAT, GSH-Px, superoxide anion scavenging ability, and hydroxyl radical scavenging ability in the liver, kidney, and testis (P < 0.05). Moreover, MDA levels were significantly reduced in plasma, liver, kidney, and testis (P < 0.01), compared to the control group. Furthermore, the dietary administration of Se significantly increased TrxR2 and GPx4 mRNA levels in kidney and testis, and ID1 mRNA levels in liver and kidney. Most of the antioxidant parameters and selenoprotein-related gene expressions significantly increased, and MDA significantly decreased at dietary supplementation with 0.5 mg/kg Se. Whereas a higher dose of Se level (1 or 2 mg/kg) inhibited the activities of some of the antioxidant enzymes and selenoprotein-related gene expressions in selected tissues. In conclusion, dietary Se supplementation with 0.5 mg/kg significantly improved roosters' antioxidant status and selenoprotein-related gene expression in liver, kidney, and testis, while higher doses led to inhibit these; dietary Se might increase reproductive performance by enhancing their antioxidant status in roosters.

Selenomethionine protected BMECs from inflammatory injury and oxidative damage induced by Klebsiella pneumoniae by inhibiting the NF-κB and activating the Nrf2 signaling pathway.

Klebsiella pneumoniae (K. pneumoniae) is one of the main environmental pathogens causing bovine mastitis. The incidence of bovine mastitis caused by K. pneumoniae is increasing worldwide. Selenium is an essential trace element that has multiple physiological functions, such as antioxidant and anti-inflammatory activities. Therefore, this study aimed to verify whether selenomethionine (SeMet) could contribute to alleviating the inflammatory injury and oxidative damage induced by K. pneumoniae. Bovine mammary epithelial cells were cultured in vitro and pretreated with 4 µM SeMet before being infected with K. pneumoniae. Western blot analysis was used to detect the expression of the related proteins in the NF-κB and Nrf2 signaling pathways. The gene expression levels of IL-1ß, IL-6, IL-8, TNF-α, Nrf2, Keap1, NQO-1 and HO-1 were detected using RT-qPCR. The levels of MDA, GSH-PX, SOD, CAT and T-AOC were detected by commercial assay kits. Flow cytometry was used to determine the level of intracellular ROS, and immunofluorescence was used to detect the nuclear localization of Nrf2 protein. Briefly, SeMet downregulated the phosphorylation levels of IκBα and p65 proteins and the gene expression levels of IL-1ß, IL-6, IL-8 and TNF-α were also decreased. Moreover, the protein and gene expression levels of Nrf2, NQO-1 and HO-1 were upregulated, and the nuclear expression of Nrf2 protein was also promoted, which enhanced the activity of antioxidant enzymes. In conclusion, SeMet protected BMECs from inflammatory injury and oxidative stress induced by K. pneumoniae by inhibiting the NF-κB and activating the Nrf2 signaling pathway.

Effect of Dietary Methylseleninic Acid and Se-Methylselenocysteine on Carcinogen-Induced, Androgen-Promoted Prostate Carcinogenesis in Rats.

Selenomethionine (SeMet) did not prevent prostate cancer in the SELECT trial and in two hormone-driven rat models. However, we have shown that daily oral bolus administration of next-generation selenium forms, methylseleninic acid (MSeA) and Se-methylselenocysteine (MSeC) at 3 mg Se/kg body weight, inhibits prostate carcinogenesis in the TRAMP and pten-deficient mouse models and In Vivo growth of human prostate cancer cells. Here, we determined whether these Se forms prevent prostate cancer in a chemically induced-androgen promoted carcinogenesis rat model in which SeMet was not preventive. WU rats were treated with methylnitrosourea, and one week later, slow-release testosterone implants when they were randomized to groups fed AIN-93M diet supplemented with 3 ppm selenium as MSeA or MSeC or control diet. Mean survival, tumor incidence in all accessory sex glands combined (dorsolateral and anterior prostate plus seminal vesicle) and the incidence of tumors confined to dorsolateral and/or anterior prostate were not statistically significantly different among the groups. Thus, MSeA and MSeC feeding was not preventive in this model. The contrast with the inhibitory effects of MSeA and MSeC in mouse models may be due to differences in carcinogenic mechanisms, selenium dosage, delivery mode, and pharmacokinetics or fundamental rat-mouse differences in selenium metabolism.

Production of selenomethionine labeled polyglycine hydrolases in Pichia pastoris.

Producing recombinant proteins with incorporated selenomethionine (SeMet) facilitates solving X-ray crystallographic structures of novel proteins. Production of SeMet labeled proteins in the yeast Pichia pastoris (syn. Komagataella phaffii) is difficult because SeMet is mildly toxic, reducing protein expression levels. To counteract this yield loss for a novel protease, Epicoccum sorghi chitinase modifying protein (Es-cmp), a novel disease promoting protease secreted by these plant pathogenic fungi, we isolated a yeast strain that secreted more protein. By comparing the expression level of 48 strains we isolated one that produced significantly more protein. This strain was found to be gene dosed, having four copies of the expression cassette. After optimization the strain produced Es-cmp in defined media with SeMet at levels nearly equal to that of the original strain in complex media. Also, we produced SeMet labeled protein for a homologous protease from the fungus Fusarium vanettenii, Fvan-cmp, by directly selecting a gene dosed strain on agar plates with increased zeocin. Linearization of plasmid with PmeI before electroporation led to high numbers of 1 mg/mL zeocin resistant clones with significantly increased expression compared to those selected on 0.1 mg/mL. The gene dosed strains expressing Es-cmp and Fvan-cmp allowed production of 8.5 and 16.8 mg of SeMet labeled protein from 500 mL shake flask cultures. The results demonstrate that selection of P. pastoris expression strains by plating after transformation on agar with 1 mg/mL zeocin rather than the standard 0.1 mg/mL directly selects gene dosed strains that can facilitate production of selenomethionine labeled proteins.

Selenium-containing Peptides and their Biological Applications.

Selenium (Se) has been known for its beneficial biological roles for several years, but interest in this trace element has seen a significant increase in the past couple of decades. It has been reported to be a part of important bioactive organic compounds, such as selenoproteins and amino acids, including selenocysteine (SeCys), selenomethionine (SeMet), selenazolidine (SeAzo), and selenoneine. The traditional Se supplementations (primarily as selenite and selenomethionine), though have been shown to carry some benefits, also have associated toxicities, thereby paving the way for the organoselenium compounds, especially the selenoproteins and peptides (SePs/SePPs) that offer several health benefits beyond fulfilling the elementary nutritional Se needs. This review aims to showcase the applications of selenium-containing peptides that have been reported in recent decades. This article summarizes their bioactivities, including neuroprotective, antiinflammatory, anticancer, antioxidant, hepatoprotective, and immunomodulatory roles. This will offer the readers a sneak peek into the current advancements to invoke further developments in this emerging research area.

Protective Effect of Selenomethionine on T-2 Toxin-Induced Rabbit Immunotoxicity.

T-2 toxin is a trichothecene mycotoxin produced by fusarium species, which is mainly prevalent in grain and livestock feed. One of the main effects of this toxin is immunodepression. Previous studies have shown that T-2 toxin can cause damage to immune organs and impaired immune function in animals. However, selenomethionine (SeMet) as an organic selenium source can not only promote the growth and development of the body but also effectively improve the body's immune function. In this study, rabbits were exposed to 0.4-mg/kg T-2 toxin, and abnormal blood routine indicators were found in the rabbits. HE staining also showed obvious lesions in the spleen and thymus tissue structures, accompanied by a large number of bleeding points. In addition, rabbits showed strong oxidative stress and inflammatory response after T-2 toxin action. 0.2 mg/kg, 0.4 mg/kg, and 0.6 mg/kg organic selenium were added to the feed. However, it was found that 0.2 mg/kg selenium can effectively improve the abnormal changes of blood routine and spleen and thymus tissue of rabbits. On the other hand, it can significantly increase the expression of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), and total antioxidant capacity (T-AOC) in the spleen and thymus, and downregulate the expression of reactive oxygen species (ROS) and malondialdehyde (MDA). In addition, inflammatory factors interleukin-1 beta (IL-1ß) and interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) in blood were also significantly inhibited; the expression of proliferating cell nuclear antigen (PCNA) in the spleen and thymus was also significantly increased after low-dose selenium treatment. Surprisingly, 0.4 mg/kg and 0.6 mg/kg of selenium did not effectively alleviate the immunotoxic effects caused by T-2 toxin, and cause damage to a certain extent. In summary, our results show that 0.2 mg/kg of SeMet can effectively alleviate the immunotoxicity caused by T-2 toxin. Selenium may protect rabbits from T-2 toxin by improving its antioxidant and anti-inflammatory capabilities.

Long-term effect of parental selenium supplementation on the one-carbon metabolism in rainbow trout (Oncorhynchus mykiss) fry exposed to hypoxic stress.

This study evaluated how different forms of selenium (Se) supplementation into rainbow trout broodstock diets modified the one-carbon metabolism of the progeny after the beginning of exogenous feeding and followed by hypoxia challenge. The progeny of three groups of rainbow trout broodstock fed either a control diet (Se level: 0·3 µg/g) or a diet supplemented with inorganic sodium selenite (Se level: 0·6 µg/g) or organic hydroxy-selenomethionine (Se level: 0·6 µg/g) was cross-fed with diets of similar Se composition for 11 weeks. Offspring were sampled either before or after being subjected to an acute hypoxic stress (1·7 mg/l dissolved oxygen) for 30 min. In normoxic fry, parental Se supplementation allowed higher glutathione levels compared with fry originating from parents fed the control diet. Parental hydroxy-selenomethionine treatment also increased cysteine and cysteinyl-glycine concentrations in fry. Dietary Se supplementation decreased glutamate-cysteine ligase (cgl) mRNA levels. Hydroxy-selenomethionine feeding also lowered the levels of some essential free amino acids in muscle tissue. Supplementation of organic Se to parents and fry reduced betaine-homocysteine S-methyltransferase (bhmt) expression in fry. The hypoxic stress decreased whole-body homocysteine, cysteine, cysteinyl-glycine and glutathione levels. Together with the higher mRNA levels of cystathionine beta-synthase (cbs), a transsulphuration enzyme, this suggests that under hypoxia, glutathione synthesis through transsulphuration might have been impaired by depletion of a glutathione precursor. In stressed fry, S-adenosylmethionine levels were significantly decreased, but S-adenosylhomocysteine remained stable. Decreased bhmt and adenosylmethionine decarboxylase 1a (amd1a) mRNA levels in stressed fry suggest a nutritional programming by parental Se also on methionine metabolism of rainbow trout.

Propargylic Se-adenosyl-l-selenomethionine: A Chemical Tool for Methylome Analysis.

Devising synthetic strategies to construct a covalent bond is a common research topic among synthetic chemists. A key driver of success is the high tunability of the conditions, including catalysts, reagents, solvents, and reaction temperature. Such flexibility of synthetic operations has allowed for the rapid exploration of a myriad of artificial synthetic transformations in recent decades. However, if we turn our attention to chemical reactions controlled in living cells, the situation is quite different; the number of hit substrates for the reaction-type is relatively small, while the crowded environment is chemically complex and inflexible to control.A specific objective of this Account is to introduce our chemical methylome analysis as an example of bridging the gap between chemistry and biology. Protein methylation, catalyzed by protein methyltransferases (MTases) using S-adenosyl-l-methionine (SAM or AdoMet) as a methyl donor, is a simple but important post-translational covalent modification. We aim to efficiently identify MTase substrates and methylation sites using activity-based protein profiling (ABPP) with propargylic Se-adenosyl-l-selenomethionine (ProSeAM, also called SeAdoYn). Specifically, we draw heavily from quantitative proteomics that yields information about the differences between two samples utilizing LC-MS/MS analysis. By exploiting the use of ProSeAM, we have prepared the requisite two samples for quantitative methylome analysis. The structural difference between ProSeAM and the parent SAM is so small that the quantity of modification of the protein substrate with this artificial cofactor reflects, to a large extent, levels of activity of the MTase of interest with SAM. First, we identified that the addition of exogenous recombinant MTase (methylation accel), a natural catalyst, enhances the generation of the corresponding propargylated product even in the cell lysate. Then, we applied the principle to isotope label-free quantification with HEK293T cell lysates. By comparing the intensity of LC-MS/MS signals in the absence and presence of the MTase, we have successfully correlated the MTase substrates. We have currently applied the concept to the stable isotope label-based quantification, SILAC (stable isotope labeling by amino acids in cell culture). The strategy merging ProSeAM/MTase/SILAC (PMS) is uniquely versatile and programmable. We can choose suitable cell lines, subcellular fractions (i.e.; whole lysate or mitochondria), and genotypes as required. In particular, we would like to emphasize that the use of cell lysates derived from disease-associated MTase knockouts (KOs) holds vast potential to discover functionally unknown but biologically important methylation events. By adding ProSeAM and a recombinant MTase to the lysates derived from KO cells, we successfully characterized unprecedented nonhistone substrates of several MTases. Furthermore, this chemoproteomic procedure can be applied to explore MTase inhibitors (methylation brake). The combined strategy with ProSeAM/inhibitor/SILAC (PIS) offers intriguing opportunities to explore nonhistone methylation inhibitors.Considering that SAM is the second most widely used enzyme-substrate following ATP, the interdisciplinary research between chemistry and biology using SAM analogs has a potentially huge impact on a wide range of research fields associated with biological methylation. We hope that this Account will help to further delineate the biological function of this important class of enzymatic reaction.

Peanut selenium distribution, concentration, speciation, and effects on proteins after exogenous selenium biofortification.

Fortification of Se is vital importance for both nutritional demand and prevention of Se-deficiency-related diseases. To better understand t selenium distribution, concentration, speciation, its effects on proteins, and cytotoxic activity, the biofortification of exogenous Se in peanut was conducted in this study. Our data have shown that foliar spraying of Se-riched fertilizer was more efficient for biotransformation of inorganic Se to organic Se by peanut plant. Besides, the Se content in peanut was increased in a dose-dependent manner. Our present study also confirmed that SeCys2, MeSeCys, and SeMet were the main Se speciation within peanut proteins. Moreover, the secondary structure and thermostability of peanut protein were altered as a result of the Se treatments, and these alterations could be attributed to the replacements of cysteine and methionine by selenocysteine and selenomethionine, respectively. The Se-enriched peanut protein could significantly inhibit the growth of Caco-2 and HepG2 in a concentration-dependent manner.

Selenomethionine as an expressible handle for bioconjugations.

Site-selective chemical bioconjugation reactions are enabling tools for the chemical biologist. Guided by a careful study of the selenomethionine (SeM) benzylation, we have refined the reaction to meet the requirements of practical protein bioconjugation. SeM is readily introduced through auxotrophic expression and exhibits unique nucleophilic properties that allow it to be selectively modified even in the presence of cysteine. The resulting benzylselenonium adduct is stable at physiological pH, is selectively labile to glutathione, and embodies a broadly tunable cleavage profile. Specifically, a 4-bromomethylphenylacetyl (BrMePAA) linker has been applied for efficient conjugation of complex organic molecules to SeM-containing proteins. This expansion of the bioconjugation toolkit has broad potential in the development of chemically enhanced proteins.

Identification of the biliary selenium metabolite and the biological significance of selenium enterohepatic circulation.

Although selenium (Se) is mainly excreted in urine, it has been reported that an unknown Se metabolite is excreted in bile. When we administered selenomethionine (SeMet), selenocyanate or selenite to rats, a common biliary selenometabolite was detected 10 min after administration. The amount of the selenometabolite originating from SeMet was less than that originating from the two inorganic Se compounds, selenocyanate and selenite, suggesting that the transformation from the methylated organic selenocompound, i.e., SeMet, was less efficient than that from the inorganic Se compounds. The common biliary selenometabolite was concretely identified as selenodiglutathione (GSSeSG) by two types of mass spectrometry, i.e., LC-inductively coupled mass spectrometry (ICP-MS) and LC-ESI-Q/TOF. The bile-drained rats had lower urinary Se levels than the sham-operated rats. In addition, the Se amounts in urine plus bile of the bile-drained rats were comparable to the Se amount in the urine of the sham-operated rats. These results suggest that the biliary selenometabolite, GSSeSG, was reabsorbed in the gut and finally excreted in urine. Enterohepatic circulation occurs to maintain Se status in the body.

Effects of different levels of dietary hydroxy-analogue of selenomethionine on growth performance, selenium deposition and antioxidant status of weaned piglets.

This study was conducted to assess the effects of the hydroxy-analogue of selenomethionine (HMSeBA) on growth performance, selenium (Se) deposition and antioxidant status of piglets. In a 28-d experiment, 252 piglets were assigned into seven treatments. These treatments were a negative control (Con-, basal diet without supplement Se), a positive control (Con+, basal diet + 0.3 mg Se from sodium selenite per kg), and five HMSeBA groups (basal diet + 0.1, 0.2, 0.3, 0.4 and 0.5 mg Se/kg from HMSeBA, respectively). Results showed that dietary HMSeBA supplementation did not affect growth performance of piglets. However, HMSeBA supplementation increased the Se concentrations in serum, liver, kidney and muscle compared with groups Con- and Con+ (p < 0.05). Compared with group Con-, supplementation with 0.2 and 0.4 mg Se from HMSeBA increased serum total antioxidant capability (T-AOC) and addition of 0.4 and 0.5 mg Se from HMSeBA increased serum glutathione peroxidase (GSH-Px) activities (p < 0.05). Compared with group Con-, the addition of 0.1, 0.2, 0.4, 0.5 mg Se from HMSeBA increased GSH-Px activities and decreased malondialdehyde (MDA) contents in the liver, and 0.3 mg Se from HMSeBA increased T-AOC and GSH-Px activities in the liver (p < 0.05). Compared with group Con+, 0.3 mg Se from HMSeBA increased serum superoxide dismutase (SOD) and hepatic T-AOC activities, and decreased the serum MDA level (p < 0.05). In general, dietary HMSeBA supplementation could improve Se deposition in serum and tissue and antioxidant capacity of piglets, suggesting that HMSeBA could be an effective Se source for piglets.

Selenium biofortification and its effect on multi-element change in Auricularia auricular.

Auricularia auricular could be useful as a candidate for human selenium supplementation. This study examined the effects of exogenous Se on the growth, yield, nutritive value, and mineral accumulation of A. auricular. Selenate or selenite (0.5-40.0 µg g-1) had no effect on mycelium morphology or the yield of fruiting bodies. In some cases, they affected the accumulation of inter-elements and significantly decreased the concentrations of copper, iron, and chromium in the Se-enriched fruiting bodies compared to that with control treatments. The polysaccharide (116.5-131.6 µg g-1) and protein (105.2-113.4 µg g-1) content in Se-enriched fruiting bodies were not significantly different from those observed in the controls (polysaccharide, 114.1 µg g-1; protein, 105.6 µg g-1). Thus, A. auricular can absorb inorganic Se from the substrate and convert it to organic Se compounds (selenocystine (≥4.1%), selenomethionine (≥91.9%), and Se-methylselenocysteine (≥2.3%)).

Dietary selenomethionine increases antioxidant capacity of geese by improving glutathione and thioredoxin systems.

A total of 200 healthy 28-day-old male Jiangnan White geese were used to investigate the free radical scavenging ability, reduced glutathione (GSH) and thioredoxin systems, and the concentrations of reactive oxygen metabolites (ROM), malondialdehyde (MDA), and protein carbonyl (PC) in geese fed diets with organic selenium (Se) (Selenomethionine, SeMet) and inorganic Se (sodium selenite, SS). All geese were randomly allotted into 4 groups with 5 replicates of 10 geese each, and received basal diet supplemented with 0.3 mg Se/kg SS, 0.2, 0.3 and 0.4 mg Se/kg SeMet until 70 D of age, respectively. Geese in the SS and SeMet groups exhibited similar growth performance. Diet with SeMet increased the scavenging abilities of 2,2-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt free radical (ABTS•+, P < 0.001) and superoxide radical (O2-•, P = 0.002) in the serum of geese, as well as the scavenging abilities of ABTS•+ (P = 0.023), hydroxyl radical (P = 0.009) and O2-• (P = 0.019) in the liver of geese. Compared to the SS group, SeMet increased hepatic GSH concentration (P = 0.002), the activities of glutathione peroxidase (P = 0.031), γ-glutamate cysteine ligase (P < 0.001), and thioredoxin reductase (P < 0.001), and decreased the concentrations of ROM, MDA, and PC in the serum and liver of geese (P < 0.05). In conclusion, dietary SeMet inclusion would be more effective than SS in increasing the antioxidant capacity of geese, possibly by improving GSH and thioredoxin systems, and 0.2 mg Se/kg SeMet in goose diet is recommended.

Methioninase Gene Therapy.

Recombinant methioninase (rMETase) derived from Pseudomonas putida targets the elevated methionine (MET) requirement of cancer cells (methionine dependence) and has shown efficacy against a variety of cancer types in mouse models. To enhance the efficacy of rMETase, we constructed the pLGFP-METSN retrovirus encoding the P. putida methioninase (METase) gene fused with the green fluorescent protein (GFP) gene. pLGFP-METSN or control vector pLGFPSN was introduced into the human lung cancer cell line H460. The retrovirus-mediated METase gene transfer decreased the intracellular MET level of the cancer cells and consequently enhanced the efficacy of treatment with the rMETase protein. The rMETase gene was introduced into an adenovirus. rAd-METase transduction of human OVACAR-8 ovarian cancer cells and human fibrosarcoma HT1080 cells in vitro and in vivo resulted in high levels of METase expression up to 10% or more of the total protein of the cells, depending on the multiplicity of infection. The combination of rAd-METase and rMETase was synergistic to kill these cells. Normal fibroblasts, on the other hand, appeared relatively resistant to the METase gene in the presence of rMETase. Adenoviral METase-transduced cancer cells were used in combination with selenomethionine, releasing highly toxic methylselenol, which killed both the cancer cells containing the METase gene and bystanders. Methylselenol damaged the mitochondria via oxidative stress and caused cytochrome c release into the cytosol, thereby activating the caspase cascade and cancer-cell apoptosis. Adenoviral METase-gene/SeMET treatment also inhibited tumor growth in rodents and significantly prolonged their survival. AdMETase/SeMET therapy was effective against Bcl-2-overproducing A549 lung cancer cells, which were resistant to staurosporine-induced apoptosis, with a strong bystander effect. The combination of Ad-METase/SeMET and doxorubicin (DOX) delayed the growth of the H460 human lung cancer, growing subcutaneously in nude mice. These results demonstrate the potential of methionine restriction (MR) for cancer treatment.

Direct Imaging of Protein-Specific Methylation in Mammalian Cells.

Abundant post-translational modification through methylation alters the function, stability, and/or localization of a protein. Malfunctions in post-translational modification are associated with severe diseases. To unravel protein methylation sites and their biological functions, chemical methylation reporters have been developed. However, until now, their usage was limited to cell lysates. Herein, we present the first generally applicable approach for imaging methylation of individual proteins in human cells, which is based on a combination of chemical reporter strategies, bioorthogonal ligation reactions, and FRET detected by means of fluorescence lifetime imaging microscopy. Through this approach, methylation of histone 4 and the non-histone proteins tumor suppressor p53, kinase Akt1, and transcription factor Foxo1 in two human cell lines has been successfully imaged. To further demonstrate its potential, the localization-dependent methylation state of Foxo1 in the cellular context has been visualized.