Multi-dimensional HPLC/MS of the nucleolar proteome using HPLC-chip/MS

The proteome of the human nucleolus was investigated in a single analysis using off-line strong cation exchange chromatography and microfraction collection combined with HPLC-chip/MS. The analysis was conducted either as a 1-D workflow with HPLC-chip alone or as a 2-D workflow. Two hundred and six unique proteins were identified in the International Protein Index human database corresponding to 2024 unique tryptic peptides identified in the 2-D analysis. In contrast, only 34 proteins and 151 corresponding tryptic peptides were found by applying a 1-D separation strategy. This clearly indicated that the complexity of the samples required the combination of more than one orthogonal separation technique. Stringent database search criteria, including reversal of sequences and therefore better exclusion of false-positive identifications, were applied for reliable protein identification.     

The stability of SiC coating and SiO2/SiC multilayer on the surface of graphite for HTGRs at normal service condition

The stability of SiC coating in helium with a low concentration of O₂, CO₂, and H₂O is a key factor for their application in improvement of oxidation resistance of graphite for high temperature gas-cooled reactors (HTGRs). Through thermodynamic analysis, it is found that the influence factor controlling the critical temperature of passive oxidation for SiC is partial pressure of active gas in helium; the critical temperature of passive oxidation for SiC increases with the partial pressure of O₂, CO₂, and H₂O, SiC is prone to undergo active oxidation in He–CO₂ and He–H₂O system. SiO₂/SiC multilayer coating can improve the oxidation resistance of graphite at higher temperature than SiC coating does under normal operation condition for HTGRs.     

Effect of preadsorbed water on the adsorption of p-xylene and m-xylene mixtures on BaX and BaY zeolites

Adsorption of an equimolar p-xylene/m-xylene mixture on partially hydrated barium-exchanged X and Y zeolites is studied at 423 K in the pressure range 10(-2)-8 hPa by differential calorimetry coupled with manometry and chromatography. Results are consistent with structural studies and Monte Carlo simulations of the literature. The presence of preadsorbed water in supercages increases the adsorption selectivity toward p-xylene to the detriment of the adsorption capacity and the adsorption affinity, as indicated by a sharp decrease of the Henry constants. However, the coadsorption heats at zero filling are not influenced by the presence of water. Therefore, entropic effects seem to play an important role in the coadsorption process. Two adsorption sites whose energy differs are identified by calorimetry. The more energetic site could correspond to the p-xylene or m-xylene molecule in interaction with the compensation cation located in sites II in the supercage, the less energetic to the adsorption of p-xylene molecule in the 12-ring window joining two supercages. The presence of this second site for p-xylene could be at the origin of the selectivity.     

A novel diastereoselective synthesis of (Z)-fluoroalkenes via a Nozaki-Hiyama-Kishi-type reaction

A highly diastereoselective and straightforward synthesis for (Z)-2-fluoroallylic alcohols via a Nozaki-Hiyama-Kishi-type reaction with the corresponding bromofluoroolefins was developed, providing an easy access to highly interesting fluorinated synthons.     

Enhanced stereoselectivity in internucleotidic bond formation by the use of the chiral ribose moiety of thymidine

This paper deals with the synthesis of new cyclic thymidine 3'-phosphoramidite building blocks having a covalent linker between the trityl type 5'-hydroxyl protecting group and the phosphorus atom attached to the 3'-hydroxyl group of thymidine. The ring structures were designed to reduce the conformational freedom around the phosphorus center so that the stereoselectivity in the internucleotide linkage formation would be improved. The linkers were also designed to be removed readily by treatment with aqueous ammonia. These building blocks were synthesized in good yield by one-pot cyclization of the diol precursors with dichloro(N,N-diisopropylamino)phosphine, despite their large-membered ring. Various activators having 1H-tetrazole, imidazole, and triazole structures were investigated to find the best selectivity in the synthesis of thymidylyl(5'-3')thymidine phosphorothioate. It turned out that our cyclic phosphoramidites gave preferentially the R(p) diastereoisomer in high coupling yield applicable to the solid-phase synthesis of oligodeoxynucleotides. It should be noted that high stereoselectivity was achieved without any chiral sources other than the 2'-deoxyribose moiety itself. The mechanistic studies revealed the importance of the steric bulk and the acidity of the activators. It was also found that the steric bulk of the alcoholic nucleophile was an important factor that determined the stereoselectivity in our systems.     

Étude des conditions d'accès au tétrazolo-1,2,3,4-[5′,1′:6,1] triazino-1,2,4-[4,5-b]indazole et au thioxo-3 triazolo-1,2,4-[2′,1′:6,1] triazino-1,2,4-[4,5-b]indazole

The triazinoindazolones 8 and 9 were synthesized by rearrangement of the oxadiazoles 5 and 6 or by cyclising the indazolic ethoxymethylidene-hydrazides 2 and 3. The triazinoindazolethiones 17 and 18 which were obtained by thionation of the triazinoindazolones 8 and 9 constitute the intermediates to the hydrazinoindazoles 19 and 20. The tetrrazolotriazinoindazole 21 was obtained by cyclising the hydrazinoindazole 19. The potassium indazolyldithiocarbazate 12 has been used for the synthesis of 4-thioxotriazinoindazolone 10 and 4-amino-3-thioxotriazolylindazole 15. The cyclisation of the latter gave the 3-thioxotriazinoindazolone 16. The structure of the derivatives was confirmed by ¹H-nmr.     

Hydrogen/deuterium exchange for higher specificity of protein identification by peptide mass fingerprinting

Genome sequencing projects produce large amounts of information that could be translated into potential protein sequences. Such amounts of material continuously increase protein database sizes. At present, 22 times more protein sequences are available in the SWISS-PROT and TrEMBL databases than 8 years ago in SWISS-PROT. One of the methods of choice for protein identification makes use of specific endoproteolytic cleavage followed by matrix-assisted laser desorption/ionisation mass spectrometric (MALDI-MS) analysis of the digested product. Since 1993, when this technique was first demonstrated, the conditions required for a correct identification have changed dramatically. Whilst 4-5 peptides with an uncertainty of 2-3 Da were sufficient for a correct identification in 1993, 10-13 peptides with less than 60 ppm mass error are now required for human and E. coli proteins. This evolution is directly related to the continuous increase in protein database sizes, which causes an increase in the number of false positive matches in identification results. Use of an information complement deduced from the primary protein sequence, in the process of identification by peptide mass fingerprints, can help to increase confidence in the identification results. In this article, we propose the exchange of labile hydrogen atoms with deuterium atoms to provide an alternative information complement. The exchange reaction with optimised techniques has shown an average 95% of hydrogen/deuterium (H/D) exchange on tryptic peptides. This level of exchange was sufficient to single out one or more peptides from a list of potential candidate proteins due to the dependence of H/D exchange on the peptide primary structure. This technique also has clear advantages in the identification of small proteins where direct protein identification is impaired by the limited number of endoproteolytic peptides. Then, information related to primary sequence obtained with this technique could help to identify proteins with high confidence without any expensive tandem mass spectrometry instruments.     

Synthèse et détermination de la structure de la 3H-(indazolyl-3′)-5 oxadiazol-1,3,4-one-2 par RMN 13C et spectrometrie de masse

The 3H-5-(3′-indazolyl)-1,3,4-oxadiazol-2-one was obtained by cyclising the hydrazide of the 3-indazolecarboxylic acid with carbonyl chloride. The structure was assigned by ¹³C nmr and mass spectrometry comparing the 2H, 3H-triazino-1,2,3[4,5-b]indazoledione-1,4. Rearrangement into 3H-5-(3′-indazolyl)-1,3,4-oxadiazol-2-one precedes the fragmentation of 2H,3H-triazino-1,2,4-[4,5-b]indazoledione-1,4 upon electron-impact.     

Pyrrolo[1,2-a]thiéno[2,3-e]pyrazine. Etude chimique de la dihydro-4,5 oxo-5 pyrrolo[1,2-a]thiéno[2,3-e]pyrazine

The 4,5-dihydropyrrolo[1,2-e]pyrazin-5-one (1) exhibits lactam-lactim tautomerism. N-Electrophilic substitutions could be achieved via the intermediate of the substitutions could be achieved via the intermediate of the sodium salt la. The 5-chloropyrazine 11 obtained by chlorodehydroxylation of pyrazinone 1 allowed nucleophilic substitution. Thionation of the pyrazinone 1 afforded the thioxopyrazine 27 wich gave studied and the tautomerism of the hydrazinopyrazine 14 and its derivatives are discussed in terms of hydrazone and azine structures