Heterocyclic and Alkyne Terpenoids by Terpene Synthase-Mediated Biotransformation of Non-Natural Prenyl Diphosphates: Access to New Fragrances and Probes.

Two terpene cyclases were used as biocatalytic tool, namely, limonene synthase from Cannabis sativa (CLS) and 5-epi-aristolochene synthase (TEAS) from Nicotiana tabacum. They showed significant substrate flexibility towards non-natural prenyl diphosphates to form novel terpenoids, including core oxa- and thia-heterocycles and alkyne-modified terpenoids. We elucidated the structures of five novel monoterpene-analogues and a known sesquiterpene-analogue. These results reflected the terpene synthases' ability and promiscuity to broaden the pool of terpenoids with structurally complex analogues. Docking studies highlight an on-off conversion of the unnatural substrates.

Redox metabolism: ROS as specific molecular regulators of cell signaling and function.

Redox reactions are intrinsically linked to energy metabolism. Therefore, redox processes are indispensable for organismal physiology and life itself. The term reactive oxygen species (ROS) describes a set of distinct molecular oxygen derivatives produced during normal aerobic metabolism. Multiple ROS-generating and ROS-eliminating systems actively maintain the intracellular redox state, which serves to mediate redox signaling and regulate cellular functions. ROS, in particular hydrogen peroxide (H2O2), are able to reversibly oxidize critical, redox-sensitive cysteine residues on target proteins. These oxidative post-translational modifications (PTMs) can control the biological activity of numerous enzymes and transcription factors (TFs), as well as their cellular localization or interactions with binding partners. In this review, we describe the diverse roles of redox regulation in the context of physiological cellular metabolism and provide insights into the pathophysiology of diseases when redox homeostasis is dysregulated.

Redox regulation of the insulin signalling pathway.

The peptide hormone insulin is a key regulator of energy metabolism, proliferation and survival. Binding of insulin to its receptor activates the PI3K/AKT signalling pathway, which mediates fundamental cellular responses. Oxidants, in particular H2O2, have been recognised as insulin-mimetics. Treatment of cells with insulin leads to increased intracellular H2O2 levels affecting the activity of downstream signalling components, thereby amplifying insulin-mediated signal transduction. Specific molecular targets of insulin-stimulated H2O2 include phosphatases and kinases, whose activity can be altered via redox modifications of critical cysteine residues. Over the past decades, several of these redox-sensitive cysteines have been identified and their impact on insulin signalling evaluated. The aim of this review is to summarise the current knowledge on the redox regulation of the insulin signalling pathway.

Fluorescent spherical mesoporous silica nanoparticles loaded with emodin: Synthesis, cellular uptake and anticancer activity.

The natural product emodin (EO) exhibits anti-inflammatory, antiangiogenesis and antineoplastic properties in vitro and in vivo. Due to its biological properties as well as its fluorescence, EO can be useful in pharmacology and pharmacokinetics. To enhance its selectivity to cancer cells, EO was loaded into non-fluorescent and novel fluorescent spherical mesoporous nanoparticles bearing N-methyl isatoic anhydride (SNM~M) or lissamine rhodamine B sulfonyl moieties (SNM~L). The propylamine functionalized mesoporous silica nanomaterial (SNM) were characterized by powder X-ray diffraction (XRD), nitrogen gas sorption, scanning electron microscopy (SEM), transmission electron microscopy (TEM), fluorescence spectroscopy, thermogravimetric analysis (TGA) and UV spectroscopy. The cytotoxicity of EO-loaded nanoparticles was tested against the human colon carcinoma cell line HT-29. Non-loaded SNM did not affect cell proliferation, whereas those loaded with EO were at least as efficient as EO alone. It could be shown by fluorescence microscopy that the uptake of silica nanomaterial by the tumor cells occurred within 2 h and the release of EO occurred within 48 h of treatment. Flow cytometry and Western blot analysis showed that SNM containing EO induced apoptosis in HT-29 cells.

Redox signalling and ageing: insights from Drosophila.

Ageing and age-related diseases are major challenges for the social, economic and healthcare systems of our society. Amongst many theories, reactive oxygen species (ROS) have been implicated as a driver of the ageing process. As by-products of aerobic metabolism, ROS are able to randomly oxidise macromolecules, causing intracellular damage that accumulates over time and ultimately leads to dysfunction and cell death. However, the genetic overexpression of enzymes involved in the detoxification of ROS or treatment with antioxidants did not generally extend lifespan, prompting a re-evaluation of the causal role for ROS in ageing. More recently, ROS have emerged as key players in normal cellular signalling by oxidising redox-sensitive cysteine residues within proteins. Therefore, while high levels of ROS may be harmful and induce oxidative stress, low levels of ROS may actually be beneficial as mediators of redox signalling. In this context, enhancing ROS production in model organisms can extend lifespan, with biological effects dependent on the site, levels, and specific species of ROS. In this review, we examine the role of ROS in ageing, with a particular focus on the importance of the fruit fly Drosophila as a powerful model system to study redox processes in vivo.

Sugar-Induced Obesity and Insulin Resistance Are Uncoupled from Shortened Survival in Drosophila.

High-sugar diets cause thirst, obesity, and metabolic dysregulation, leading to diseases including type 2 diabetes and shortened lifespan. However, the impact of obesity and water imbalance on health and survival is complex and difficult to disentangle. Here, we show that high sugar induces dehydration in adult Drosophila, and water supplementation fully rescues their lifespan. Conversely, the metabolic defects are water-independent, showing uncoupling between sugar-induced obesity and insulin resistance with reduced survival in vivo. High-sugar diets promote accumulation of uric acid, an end-product of purine catabolism, and the formation of renal stones, a process aggravated by dehydration and physiological acidification. Importantly, regulating uric acid production impacts on lifespan in a water-dependent manner. Furthermore, metabolomics analysis in a human cohort reveals that dietary sugar intake strongly predicts circulating purine levels. Our model explains the pathophysiology of high-sugar diets independently of obesity and insulin resistance and highlights purine metabolism as a pro-longevity target.

Loss of epithelium-specific GPx2 results in aberrant cell fate decisions during intestinal differentiation.

The selenoprotein glutathione peroxidase 2 (GPx2) is expressed in the epithelium of the gastrointestinal tract, where it is thought to be involved in maintaining mucosal homeostasis. To gain novel insights into the role of GPx2, proteomic profiles of colonic tissues either derived from wild type (WT) or GPx2 knockout (KO) mice, maintained under selenium (Se) deficiency or adequate Se supplementation conditions were established and analyzed. Amongst the panel of differentially expressed proteins, the calcium-activated chloride channel regulator 1 (CLCA1) was significantly down-regulated in GPx2 KO versus WT mice regardless of the given Se status. Moreover, transcript levels of the isoforms CLCA2 and CLCA3 showed a similar expression pattern. In the intestine, CLCA1 is usually restricted to mucin-producing goblet cells. However, although -SeKO mice had the highest numbers of goblet cells as confirmed by significantly enhanced mRNA expression levels of the goblet cell marker mucin-2, the observed expression pattern suggests that GPx2 KO goblet cells might be limited in synthesizing CLCA1. Furthermore, transcript levels of differentiation markers such as chromogranin-1 (Chga) for enteroendocrine cells and leucine-rich repeat-containing G-protein coupled receptor 5 (Lgr5) for stem cells were also downregulated in GPx2 KO mice. Moreover, this was accompanied by a downregulation of the mRNA expression levels of the intestinal hormones glucagon-like peptide 1 (Glp1), ghrelin (Ghrl) and somatostatin (Sst). Thus, it seems that GPx2 might be important for the modulation of cell fate decisions in the murine intestinal epithelium.

Modulation of MHC class I surface expression in B16F10 melanoma cells by methylseleninic acid.

The essential trace element selenium (Se) might play a role in cancer prevention as well as for cancer therapy. Its metabolite methylselenol is able to kill cells through distinct mechanisms including induction of reactive oxygen species, DNA damage and apoptosis. Since methylselenol affects innate immune responses by modulating the expression of NKG2D ligands, the aim of this study was to determine whether the methylselenol generating compound methylseleninic acid (MSA) influences the expression of the MHC class I surface antigens and growth properties thereby reverting immune escape. Treatment of B16F10 melanoma cells expressing low basal MHC class I surface antigens with dimethyldiselenide (DMDSe) and MSA, but not with selenomethionine and selenite resulted in a dose-dependent upregulation of MHC class I cell surface antigens. This was due to a transcriptional upregulation of some major components of the antigen processing machinery (APM) and the interferon (IFN) signaling pathway and accompanied by a reduced migration of B16F10 melanoma cells in the presence of MSA. Comparative "ome"-based profilings of untreated and MSA-treated melanoma cells linked the anti-oxidative response system with MHC class I antigen processing. Since MSA treatment enhanced MHC class I surface expression also on different human tumors cell lines, MSA might affect the malignant phenotype of various tumor cells by restoring MHC class I APM component expression due to an altered redox status and by partially mimicking IFN-gamma signaling thereby providing a novel mechanism for the chemotherapeutic potential of methylselenol generating Se compounds.

Altered protein expression pattern in colon tissue of mice upon supplementation with distinct selenium compounds.

The essential trace element selenium (Se) is controversially discussed concerning its role in health and disease. Its various physiological functions are largely mediated by Se incorporation in the catalytic center of selenoproteins. In order to gain insights into the impact of Se deficiency and of supplementation with different Se compounds (selenite, selenate, selenomethionine) at defined concentrations (recommended, 150 µg/kg diet; excessive, 750 µg/kg diet) in murine colon tissues, a 20-week feeding experiment was performed followed by analysis of the protein expression pattern of colon tissue specimens by 2D-DIGE and MALDI-TOF MS. Using this approach, 24 protein spots were identified to be significantly regulated by the different Se compounds. These included the antioxidant enzyme peroxiredoxin-5 (PRDX5), proteins with binding capabilities, such as cofilin-1 (COF1), calmodulin, and annexin A2 (ANXA2), and proteins involved in catalytic processes, such as 6-phosphogluconate dehydrogenase (6PGD). Furthermore, the Se compounds demonstrated a differential impact on the expression of the identified proteins. Selected target structures were validated by qPCR and Western blot which mainly confirmed the proteomic profiling data. Thus, novel Se-regulated proteins in colon tissues have been identified, which expand our understanding of the physiologic role of Se in colon tissue.

Methionine and seleno-methionine type peptide and peptoid building blocks synthesized by five-component five-center reactions.

A first example of 5-component 5-center reactions with isonitriles [Ugi-5CRs] is described. The extended Ugi type reactions involve selenoaldehydes as well as ammonia, both challenging reactants in multicomponent (MCR) systems, to generate methionine and Se-methionine moieties and derivatives as protected building blocks or for direct ligation in peptides or peptoids. The peptoid/peptide building blocks proved to be non-cytotoxic but increased the expression of genes encoding for stress protective selenoproteins (Gpx1).

Individual effects of different selenocompounds on the hepatic proteome and energy metabolism of mice.

BACKGROUND: Selenium (Se) exerts its biological activity largely via selenoproteins, which are key enzymes for maintaining the cellular redox homeostasis. However, besides these beneficial effects there is also evidence that an oversupply of Se might increase the risk towards developing metabolic disorders. To address this in more detail, we directly compared effects of feeding distinct Se compounds and concentrations on hepatic metabolism and expression profiles of mice. METHODS: Male C57BL6/J mice received either a selenium-deficient diet or diets enriched with adequate or high doses of selenite, selenate or selenomethionine for 20weeks. Subsequently, metabolic parameters, enzymatic activities and expression levels of hepatic selenoproteins, Nrf2 targets, and additional redox-sensitive proteins were analyzed. Furthermore, 2D-DIGE-based proteomic profiling revealed Se compound-specific differentially expressed proteins. RESULTS: Whereas heterogeneous effects between high concentrations of the Se compounds were observed with regard to body weight and metabolic activities, selenoproteins were only marginally increased by high Se concentrations in comparison to the respective adequate feeding. In particular the high-SeMet group showed a unique response compromising higher hepatic Se levels in comparison to all other groups. Accordingly, hepatic glutathione (GSH) levels, glutathione S-transferase (GST) activity, and GSTpi1 expression were comparably high in the high-SeMet and Se-deficient group, indicating that compound-specific effects of high doses appear to be independent of selenoproteins. CONCLUSIONS: Not only the nature, but also the concentration of Se compounds differentially affect biological processes. GENERAL SIGNIFICANCE: Thus, it is important to consider Se compound-specific effects when supplementing with selenium.

Redox proteomics: Methods for the identification and enrichment of redox-modified proteins and their applications.

PTMs are defined as covalent additions to functional groups of amino acid residues in proteins like phosphorylation, glycosylation, S-nitrosylation, acetylation, methylation, lipidation, SUMOylation as well as oxidation. Oxidation of proteins has been characterized as a double-edged sword. While oxidative modifications, in particular of cysteine residues, are widely involved in the regulation of cellular homeostasis, oxidative stress resulting in the oxidation of biomolecules along with the disruption of their biological functions can be associated with the development of diseases, such as cancer, diabetes, and neurodegenerative diseases, respectively. This is also the case for advanced glycation end products, which result from chemical reactions of keto compounds such as oxidized sugars with proteins. The role of oxidative modifications under physiological and pathophysiological conditions remains largely unknown. Recently, novel technologies have been established that allow the enrichment, identification, and characterization of specific oxidative PTMs (oxPTMs). This is essential to develop strategies to prevent and treat diseases that are associated with oxidative stress. Therefore this review will focus on (i) the methods and technologies, which are currently applied for the detection, identification, and quantification of oxPTMs including the design of high throughput approaches and (ii) the analyses of oxPTMs related to physiological and pathological conditions.

Hydrogen peroxide - production, fate and role in redox signaling of tumor cells.

Hydrogen peroxide (H2O2) is involved in various signal transduction pathways and cell fate decisions. The mechanism of the so called "redox signaling" includes the H2O2-mediated reversible oxidation of redox sensitive cysteine residues in enzymes and transcription factors thereby altering their activities. Depending on its intracellular concentration and localization, H2O2 exhibits either pro- or anti-apoptotic activities. In comparison to normal cells, cancer cells are characterized by an increased H2O2 production rate and an impaired redox balance thereby affecting the microenvironment as well as the anti-tumoral immune response. This article reviews the current knowledge about the intracellular production of H2O2 along with redox signaling pathways mediating either the growth or apoptosis of tumor cells. In addition it will be discussed how the targeting of H2O2-linked sources and/or signaling components involved in tumor progression and survival might lead to novel therapeutic targets.