Chemistry of biologically important synthetic organoselenium compounds.

Reversible inactivation of the tumor suppressor PTEN by H2O2.

Numerous studies in animal models and more recent studies in humans have demonstrated cancer chemopreventive effects with Se. There is extensive evidence that monomethylated forms of Se are critical metabolites for chemopreventive effects of Se. Induction of apoptosis in transformed cells is an important chemopreventive mechanism. Apoptosis can be triggered by micromolar levels of monomethylated forms of Se independent of DNA damage and in cells having a null p53 phenotype. Cell cycle protein kinase cdk2 and protein kinase C are strongly inhibited by various forms of Se. Inhibitory mechanisms involving modification of cysteine residues in proteins by Se have been proposed that involve formation of Se adducts of the selenotrisulfide (S-Se-S) or selenenylsulfide (S-Se) type or catalysis of disulfide formation. Selenium may facilitate reactions of protein cysteine residues by the transient formation of more reactive S-Se intermediates. A novel chemopreventive mechanism is proposed involving Se catalysis of reversible cysteine/disulfide transformations that occur in a number of redox-regulated proteins, including transcription factors. A time-limited activation mechanism for such proteins, with deactivation facilitated by Se, would allow normalization of critical cellular processes in the early stages of transformation. There is uncertainty at the present time regarding the role of selenoproteins in chemoprevention model systems where supranutritional levels of Se are employed. Mammalian thioredoxin reductase is one selenoprotein that shows increased activity with Se supplementation in the nutritional to supranutritional range. Enhanced thioredoxin reduction could have beneficial effects in oxidative stress, but possible adverse effects are considered. Other functions of thioredoxin reductase may be relevant to cell signaling pathways. The functional status of the thioredoxin/thioredoxin reductase system during in vivo chemoprevention with Se has not been established. Some in vitro studies have shown inhibitory effects of Se on the thioredoxin system correlated with growth inhibition by Se. A potential inactivating mechanism for thioredoxin reductase or other selenoenzymes involving formation of a stable diselenide form resistant to reduction is discussed. New aspects of Se biochemistry and possible functions of new selenoproteins in chemoprevention are described.

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Selenium metabolism, selenoproteins and mechanisms of cancer prevention: complexities with thioredoxin reductase

Advances in refolding of proteins produced in E. coli

The authors were asked by the Editors of ACS Chemical Biology to write an article titled “Why Nature Chose Selenium” for the occasion of the upcoming bicentennial of the discovery of selenium by the Swedish chemist Jons Jacob Berzelius in 1817 and styled after the famous work of Frank Westheimer on the biological chemistry of phosphate [Westheimer, F. H. (1987) Why Nature Chose Phosphates, Science 235, 1173–1178]. This work gives a history of the important discoveries of the biological processes that selenium participates in, and a point-by-point comparison of the chemistry of selenium with the atom it replaces in biology, sulfur. This analysis shows that redox chemistry is the largest chemical difference between the two chalcogens. This difference is very large for both one-electron and two-electron redox reactions. Much of this difference is due to the inability of selenium to form π bonds of all types. The outer valence electrons of selenium are also more loosely held than those of sulfur. As a result,...

Why Nature Chose Selenium

The Redox Potential of Selenocystine in Unconstrained Cyclic Peptides

The methodology of regioselective cysteine pairings in synthetic multiple-cystine peptides has progressed in the past years to an efficiency that allows for at least three specific inter- and intrachain disulfide bridgings. Conformational studies on various multiple-cystine peptides like hormones, protease inhibitors, and toxins revealed that these bioactive peptides, generated by posttranslational processing of precursor proteins, are folded into miniprotein-like compact globular structures of remarkable stability. This strongly suggests protein domain or subdomain properties of these families of peptides, and thus sufficient sequence-encoded information for correct oxidative refolding under appropriate experimental conditions. From intensive research on the mechanisms and pathways of oxidative refolding of proteins in vivo and in vitro, the efficient methods have emerged for simulating nature in the regeneration of native folds not only for intact proteins, but also for protein domains and subdomains. In fact, the results obtained in the oxidative folding of excised protein fragments and of relatively low mass products of posttranslational processings show that this procedure is indeed a simple way of preparing peptides with several disulfide bonds, if optimization of reaction conditions is performed in terms of redox buffer, temperature, and additives capable of disrupting aggregates and of stabilizing nascent secondary structures. Moreover, with increased knowledge about stable, small natural cystine frameworks, their use instead of artificial templates should facilitate engineering of synthetic miniproteins with specific conformation and tailored functions.

Oxidative folding of cystine-rich peptides vs regioselective cysteine pairing strategies.

Using the Fmoc/tBu protection scheme and the p-methoxybenzyl derivative of selenocysteine, the synthesis of related peptides in the selenol-protected form could be optimized by operating the coupling steps in the absence of auxiliary bases and by reducing the piperidine treatment to the minimum time required for quantitative Fmoc cleavage. Under these conditions, β-elimination of the p-methoxybenzylselenol as the main side reaction of these syntheses, as well as epimerization of the protected selenocysteine, was largely suppressed. Conversion of the selenol- and thiol-protected bis-selenocysteine and selenocysteine, cysteine peptides into the related cyclic monomeric forms by iodine-mediated oxidation failed since a complex mixture of compounds was produced. Cleavage of the selenoether bond with mercuric acetate was found to proceed smoothly, but displacement of the heavy metal ions by treatment with excesses of thiols or hydrogen sulphide was unsuccessful since a stable Hg2+ diselenide complex was obtained. However, oxidation was achieved in good yields by the dimethylsulphoxide/trifluoroacetic acid procedure and the peptides were then used for determining the redox potential of the diselenide and selenide/sulphide bridge, respectively. © 1997 European Peptide Society and John Wiley & Sons, Ltd.

Synthesis of selenocysteine peptides and their oxidation to diselenide-bridged compounds.

Using methionine-auxotrophic Escherichia coli strains quantitative biosynthetic replacement of the methionine residues by seleno- and telluromethionine but not by methoxinine was achieved in various model proteins, clearly indicating a limited tolerance in the editing range of methionyl-tRNA synthetase. For expression of the protein variants the acetyl derivatives of the chalcogen-analogs of methionine, obtained by a new and highly efficient synthetic procedure, proved to be the ideal source in the growth media as they were found to be significantly more stable than the underivatized methionine analogs. The conformational properties in solution, the folding and unfolding parameters as well as X-ray crystallographic data confirmed the highly isomorphous character of the atomic mutants and thus the usefulness of this concept in X-ray analysis of proteins. Quantitative replacement of cysteine residues by selenocysteine has recently been achieved using cysteine-auxotrophic E. coli strains, but a selective replacement of cysteine residues by employing the natural translational machinery of selenocysteine is also conceivable. We have therefore performed a detailed study on synthetic selenocysteine-peptides in order to determine the redox potential of this cysteine analog, and thus the ability of related peptide and protein analogs to undergo the correct oxidative folding. Since the redox potential of selenocysteine was found to be significantly more reducing than that of the parent amino acid, selective formation of a diselenide bridge in presence of additional cysteine residues is highly favored as well documented in the case of the synthetic bis-selenocysteine-endothelin I analog. These results confirm that even cysteine residues may represent an interesting target for the design and expression of isomorphous heteroatomic analogs of proteins.

Dörthe Besse

Papers

The Redox Potential of Selenocystine in Unconstrained Cyclic Peptides

Oxidative folding of cystine-rich peptides vs regioselective cysteine pairing strategies.

Synthesis of selenocysteine peptides and their oxidation to diselenide-bridged compounds.

Chalcogen-analogs of amino acids. Their use in X-ray crystallographic and folding studies of peptides and proteins.

Das Redoxpotential von Selenocystin in konformativ nicht eingeschränkten cyclischen Peptiden