Self-assembly of penta-selenopeptides into amyloid fibrils.

Here, we report the synthesis of a penta-selenopeptide consisting of five benzyl protected selenocysteine residues. This selenopeptide was well characterized by both one- and two-dimensional (D) NMR spectroscopies. We find that the solution conformation is enriched with ß-sheet structures, which have a propensity to self-assemble and form amyloid fibrils.

Synthesis and semisynthesis of selenopeptides and selenoproteins.

The versatile chemistry of the genetically encoded amino acid selenocysteine (Sec) is employed in Nature to expand the reactivity of enzymes. In addition to, its role in biology, Sec is used in protein engineering to modify folding, stability, and reactivity of proteins, to introduce conjugations and to facilitate reactions. However, due to limitations related to Sec's insertion mechanism in Nature, much of the production of Sec containing peptides and proteins relies on synthesis and semisynthesis. Here, we review recent advances that have enabled the assembly of complicated selenoproteins, including novel uses of protecting groups for solid phase peptide synthesis, rapid selenoester driven chemical ligations and versatile expressed protein ligations.

From protein total synthesis to peptide transamidation and metathesis: playing with the reversibility of N,S-acyl or N,Se-acyl migration reactions.

Amide forming reactions are central to the field of peptide and protein synthesis and are considered to be poorly reversible reactions owing to the high stability of peptide bonds. One amide-forming reaction is native chemical ligation (NCL) which is driven by a sulfur to nitrogen acyl migration process from a transient thioester intermediate. However, recent studies have revealed the reversibility of the S,N-acyl shift reaction or of the related Se,N-acyl shift process using mild aqueous conditions. Such chemical processes have great potential for the chemoselective formation of peptide bonds to cysteine or selenocysteine, and open novel avenues in the field of peptide transamidation and metathesis reactions.

Synthesis of [(77)Se]-methylselenocysteine when preparing sauerkraut in the presence of [(77)Se]-selenite. Metabolic transformation of [ (77)Se]-methylselenocysteine in Wistar rats determined by LC-IDA-ICP-MS.

The use of enriched Se isotopes as tracers has provided important information on Se metabolism. However, selenium isotopes are expensive and difficult to obtain. A simple and cheap strategy based on the production of [(77)Se]-methylselenocysteine ([(77)Se]-MeSeCys) when preparing sauerkraut in the presence of [(77)Se]-selenite was developed. The resulting [(77)Se]-MeSeCys was used for evaluating the metabolic transformation of MeSeCys in Wistar rats, by feeding them with an AIN-93 M diet containing 20 % sauerkraut enriched in [(77)Se]-MeSeCys. Organs (liver, kidney, brain, testicles, and heart) were obtained after seven days of treatment and subjected to total selenium and selenium-speciation analysis by high-performance liquid chromatography coupled with isotope-dilution-analysis inductively-coupled-plasma mass spectrometry (HPLC-IDA-ICP-MS). Analysis of (77)Se-labeled organs revealed a prominent increase (more than 100 % Se-level enhancement) of selenium in the kidney and heart, whereas in the liver selenium concentration only increased by up to 20 % and it remained constant in the brain and testicles. (77)Se-enriched-sauerkraut supplementation does not alter the concentration of other essential elements in comparison to controls except for in the heart and kidney, in which selenium was positively correlated with Mg, Zn, Cu, and Mo. HPLC-ICP-MS analysis of hydrolyzed extracts after carbamidomethylation of the (77)Se-labeled organs revealed the presence of [(77)Se]-SeCys and an unknown Se-containing peak, the identity of which could not be verified by electrospray-ionization (ESI)-MS-MS. Low amounts of [(77)Se]-MeSeCys were found in (77)Se-labeled liver and kidney extracts, suggesting the incorporation of this selenium species in its intact form.

Natural and synthetic selenoproteins.

Once considered highly toxic, the element selenium is now recognized as a micronutrient essential for human health. It is inserted co-translationally into many proteins as the non-canonical amino acid selenocysteine, providing the resulting selenoprotein molecules with a range of valuable redox properties; selenocysteine is also increasingly exploited as a structural and mechanistic probe in synthetic peptides and proteins. Here we review topical investigations into the preparation and characterization of natural and artificial selenoproteins. Such molecules are uniquely suited as tools for protein chemistry and as a test bed for studying novel catalytic activities.

Large-scale, protection-free synthesis of Se-adenosyl-L-selenomethionine analogues and their application as cofactor surrogates of methyltransferases.

S-adenosyl-L-methionine (SAM) analogues have previously demonstrated their utility as chemical reporters of methyltransferases. Here we describe the facile, large-scale synthesis of Se-alkyl Se-adenosyl-L-selenomethionine (SeAM) analogues and their precursor, Se-adenosyl-L-selenohomocysteine (SeAH). Comparison of SeAM analogues with their equivalent SAM analogues suggests that sulfonium-to-selenonium substitution can enhance their compatibility with certain protein methyltransferases, favoring otherwise less reactive SAM analogues. Ready access to SeAH therefore enables further application of SeAM analogues as chemical reporters of diverse methyltransferases.

Solid-phase synthesis of reduced selenocysteine tetrapeptides and their oxidized analogs containing selenenylsulfide eight-membered rings.

A series of protected and reduced forms of model tetrapeptides that mimic the C-terminus of human thioredoxin reductases were obtained in good yields, using solid-phase peptide synthesis (SPPS). SPPS was performed on the Knorr Amide MBHA resin for Fmoc chemistry using especially protected cystein and selenocystein derivatives. All amino acids have been coupled according to the HBTU/HOBt/DIPEA method. Furthermore, the corresponding oxidized peptides containing eight-membered rings with intramolecular S-S and S-Se bridges were prepared via I[Formula: see text]/MeOH or DMSO/TFA oxidation, respectively.

Direct synthesis of sensitive selenocysteine peptides by the Ugi reaction.

Ammonia and selenoaldehydes are both problematic components in Ugi reactions. Here we report the efficient direct multicomponent synthesis of sensitive selenocysteine peptides without the use of convertible (protected) primary amines, including suitable deprotection protocols for selenols.

One-pot synthesis of highly functionalized seleno amino acid derivatives.

We herein provide a new and rapid protocol to generate derivatives of seleno amino acid, including methyl selenocysteine, selenomethionine, and selenocystine. Applying the isocyanide-based multicomponent reaction Ugi-4C-5C reaction, we show that each of the commercially available seleno amino acids are good substrate for these reactions and can be used together with complementary oxocomponents and isocyanides to generate highly diverse functionalized selenium-containing compounds. These compounds might become useful tools for applications in chemical biology to elucidate the role of selenium in biochemistry.

Synthesis of selenocysteine and its derivatives with an emphasis on selenenylsulfide (-Se-S-) formation.

A short survey of historic and current methods for the synthesis of selenocysteine, selenocystine, and derivatives and related compounds is presented, with an additional emphasis on the formation of selenocysteine-derived Se-S bridges. The majority of methods to the amino acid starts with protected and O-activated serine, but also other concepts are included such as radical or multicomponent strategies, the latter allowing also direct access to peptoids in one pot. Of special importance is the monomeric oxidative cyclization of selenocysteine-cysteine peptides to eight-membered and larger rings with a selenenylsulfide bridge, a crucial element in several selenoproteins.

Synthesis of selenocysteine and selenomethionine derivatives from sulfur-containing amino acids.

Selenium-containing amino acids have attracted increasing interest from view points of the importance as active centers of several selenoenzymes, the biological synthesis, the metabolism, and the use for structure determination of proteins. In this article, our recent progresses in the transformation from sulfur-containing amino acids to selenocysteine (SeCys) and selenomethionine (SeMet) derivatives are reviewed along with the surveys of general organic methodologies for the synthesis of SeCys and SeMet derivatives in the literature. The S-->Se modification (i.e., the chemical atomic mutation) would be a useful approach to peptide synthesis involving selenoamino acid residues.

Selenium in chemistry and biochemistry in comparison to sulfur.

What makes selenoenzymes--seen from a chemist's view--so special that they cannot be substituted by just more analogous or adapted sulfur proteins? This review compiles and compares physicochemical properties of selenium and sulfur, synthetic routes to selenocysteine (Sec) and its peptides, and comparative studies of relevant thiols and selenols and their (mixed) dichalcogens, required to understand the special role of selenium in selenoproteins on the atomic molecular level. The biochemically most relevant differences are the higher polarizability of Se- and the lower pKa of SeH. The latter has a strikingly different pH-dependence than thiols, with selenols being active at much lower pH. Finally, selected typical enzymatic mechanisms which involve selenocysteine are critically discussed, also in view of the authors' own results.

Chiral seleno-amines from indium selenolates. A straightforward synthesis of selenocysteine derivatives.

A simple and efficient procedure for the synthesis of chiral beta-seleno-amines derivatives from a one-pot indium(I) iodide-mediated aziridine ring opening with diorganoyl diselenides has been developed. As an application, the synthesis of selenocysteine and selenotreonine derivatives has been accomplished.

One pot synthesis of selenocysteine containing peptoid libraries by Ugi multicomponent reactions in water.

Selenocysteine containing peptoids and peptide-peptoid conjugates were synthesized by combinatorial Ugi-MCRs (multicomponent reactions) in water: for the first time, an acetal (selenoacetal 2a) was used in Ugi-MCR to furnish selenocysteine peptoids in one step as model compounds for selenocysteine peptides and proteins.

Dehydroalanine-containing peptides: preparation from phenylselenocysteine and utility in convergent ligation strategies.

This protocol describes the methodology for the synthesis of dehydroalanine (Dha)-containing peptides and illustrates their use in convergent ligation strategies for the preparation of peptide conjugates. A nonproteinogenic amino acid, Fmoc-Se-phenylselenocysteine (SecPh), can be prepared in high yield over four synthetic steps and be conveniently incorporated into peptides by standard solid-phase peptide synthesis techniques. Globally deprotected peptides containing phenylselenocysteine can be converted to dehydrated peptides following a chemoselective, mild oxidation with hydrogen peroxide or sodium periodate (i.e., the phenylselenocysteine side chain is converted to that of Dha). Dha residues are electrophilic handles for the preparation of glycopeptides, lipopeptides or other peptide conjugates; one such transformation will be outlined here. The preparation of Dha-containing peptides, including the synthesis of SecPh, peptide elongation and oxidative treatment of phenylselenocysteine-containing peptides can be completed by one person in approximately 3-5 weeks. However, once SecPh is in hand, the time required for the preparation of peptides is significantly shorter and comparable to that for any peptide synthesis.

Incorporation of selenocysteine into proteins using peptide ligation.

Expressed protein ligation has become a frequently used technique to insert non-standard amino acids into proteins. The technique has been adapted to insert selenocysteine residues in place of cysteine residue in proteins, taking advantage of the similarity in the chemistries of sulfur and selenium. This replacement can confer unique structural and catalytic properties to enzymes and proteins. The development of this technique also allows for naturally occurring selenoproteins to be produced semisynthetically.

Internally stabilized selenocysteine derivatives: syntheses, 77Se NMR and biomimetic studies.

Selenocystine ([Sec]2) and aryl-substituted selenocysteine (Sec) derivatives are synthesized, starting from commercially available amino acid l-serine. These compounds are characterized by a number of analytical techniques such as NMR (1H, 13C and 77Se) and TOF mass spectroscopy. This study reveals that the introduction of amino/imino substituents capable of interacting with selenium may stabilize the Sec derivatives. This study further suggests that the oxidation-elimination reactions in Sec derivatives could be used for the generation of biologically active selenols having internally stabilizing substituents.

Studies on the inhibition of human cytochromes P450 by selenocysteine Se-conjugates.

1. To investigate whether cytochrome P450 (P450) inhibition can contribute to the chemopreventive activity of selenocysteine Se-conjugates (SeCys conjugates), 21 SeCys conjugates were screened for their inhibitory potency towards seven of the most important human P450s. 2. The majority of the SeCys conjugates produced near complete inhibition of CYP1A1 at a concentration of 250 microm. The most potent inhibitor, Se-benzyl-L-selenocysteine, displayed an IC50 of 12.8 +/- 1.2 microm. CYP2C9, -2C19 and -2D6 were moderately (50-60%) inhibited by the SeCys conjugates. CYP1A2, -2E1 and -3A4 were least inhibited. 3. Studies on the susceptibility of CYP1A1 to SeCys conjugates implicated a thiol-reactive intermediate, as evidenced by reduced inhibition levels in the presence of glutathione and N-acetyl cysteine. Uncoupling of the P450-catalytic cycle was of no importance as ROS scavengers did not influence inhibition levels. 4. P450 inhibition by two physiologically relevant metabolite classes of SeCys conjugates was also studied. N-acetylation of SeCys conjugates consistently increased the inhibitory potency towards CYP1A2, -2C19, -2E1 and -3A4. Beta-lyase catalysed bioactivation of alkyl-substituted SeCys conjugates or Se-benzyl-L-selenocysteine produced little or no additional inhibition of P450 activity. For Se-phenyl-L-selenocysteine, however, significant increases in P450 inhibition were obtained by beta-lyase pre-incubation. 5. It is concluded that the potent and relatively selective CYP1A1 inhibition exerted by SeCys conjugates may contribute to their chemopreventive activity.