[Show abstract][Hide abstract] ABSTRACT: We describe a new class of silicone-containing peptide polymers obtained by a straightforward polymerization in water using tailored chlorodimethylsilyl peptide blocks as monomeric units. This general strategy is applicable to any type of peptide sequences, yielding linear or branched polymer chains composed of well-defined peptide sequences. Hybrid peptides: Silicone-containing peptide polymers are obtained from hybrid hydroxydimethylsilyl peptide blocks as monomeric units. This general strategy proceeds in water and is applicable to any type of peptide, yielding linear or branched polymer chains.
Full-text · Article · Feb 2015 · Angewandte Chemie International Edition in English
[Show abstract][Hide abstract] ABSTRACT: The detection of post-translational modifications (PTMs) of proteins is a matter of intensive research. Among all possible pitfalls that may lead to misidentifications, the chemical stability of modified peptides is scarcely questioned. Global proteomic studies devoted to protein acetylation are becoming popular. Thus, we were concerned about the intrinsic stability of O-acetylated peptides because of the O-N acyl transfer reactivity occurring when an amino moiety is present in the vicinity of the acylated hydroxyl group. Here, the behavior of isomeric O- and N-acetylated, N-terminal threonine-containing peptides was explored in a standard proteomic workflow. We demonstrated a strong chemical instability of O-acetylation, which prevents its detection.
No preview · Article · Jun 2014 · Journal of Proteomics
[Show abstract][Hide abstract] ABSTRACT: We report applications of new hybrid organic-inorganic silica based materials as laser desorption/ionization (LDI)-promoting surfaces for high-throughput identification of peptides. The driving force of our work was to design a new material composed of a conventional MALDI matrix covalently attached to silica with a high organic/inorganic ratio in order to improve the UV absorption by such LDI hybrid matrices. Amorphous CHCA-functionalized silica presenting an organic content up to 1.3 mmol g(-1) (around 40% in weight from TGA and elementary analysis measurements) gave very interesting LDI performances in terms of detection sensitivity as well as relative ionization discrepancy (spectral suppression) through the analyses of small synthetic peptide mixtures (550-1300 Da) taking CHCA and amorphous silica as model matrices for control experiments.
[Show abstract][Hide abstract] ABSTRACT: A straightforward method for the preparation of hybrid bioorganic–inorganic materials is reported. Common strategies to synthesize such promising materials require special surface modifications of silica followed by grafting of the organic moiety via chemoselective ligation. In this context, we set up a general and bottom-up strategy relying on modified peptides functionalized with a trialkoxysilane group. Used in mixtures with TEOS and a surfactant as the structure directing agent, these hybrid building blocks allow one step direct synthesis of bioorganic–inorganic hybrid materials. Two examples were chosen to demonstrate our general approach. (1) An antifouling surface was prepared by dip coating of a sol containing an antibacterial silylated peptide. (2) Organized mesoporous silica displaying a peptide catalyst in the pores was prepared in one step and tested.
[Show abstract][Hide abstract] ABSTRACT: Despite correct purity of crude peptides prepared on trityl resin by Fmoc/tBu microwave assisted solid phase peptide synthesis, surprisingly, lower yields than those expected were obtained while preparing C-terminal acid peptides. This could be explained by cyclization/cleavage through diketopiperazine formation during the second amino acid deprotection and third amino acid coupling. However, we provide here evidence that this is not the case and that this yield loss was due to high temperature promoted hydrolysis of the 2-chlorotrityl ester, yielding premature cleavage of the C-terminal acid peptides.
[Show abstract][Hide abstract] ABSTRACT: EAPB0503, belonging to the imidazo[1,2-a]quinoxaline series, is an anticancer drug with antitumoral activity against a variety of tumors. Previous studies have shown that this drug undergoes demethylation and oxygenation reactions. In this paper, liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) was employed to assess the structures of unknown oxygenated metabolites of EAPB0503. EAPB0503 and its identified demethylated metabolites, EAPB0502 and EAPB0603, were incubated with human, rat, dog and mouse liver microsomes, as well as human, rat and dog hepatocytes. After separation on a C8 analytical column with a gradient elution of acetonitrile-formate buffer, positive ESI-MS/MS experiments were performed. To facilitate metabolite identification, the detailed fragmentation pathways of the parent compounds were first studied using high-resolution MS/MS. Additional hydrogen/deuterium exchange LC-MS/MS experiments were used to support the identification and structural characterization of metabolites. Four hydroxylated metabolites were identified: M'4 and its demethylated derivative M'1 (OH in ortho position on the phenyl substituent in position 1), and M'6 and its demethylated derivative M'3 (OH on the imidazole ring at the C2 position). Three phase II metabolites (Met A, EAPB0602 glucuronide; Met B, M'4 glucuronide; Met C, EAPB0603 glucuronide) were also evidenced. Elucidation of the metabolite structures was performed by comparing the chromatographic behaviors (changes in retention times), by measuring the molecular masses (mass increment), by studying the MS(2) spectral patterns of metabolites with those of parent drugs and for M'1 and M'4 by co-analysis with synthetic standards. The results of the present study provided important structural information relating to the metabolism of EAPB0503.
Full-text · Article · Oct 2013 · Journal of pharmaceutical and biomedical analysis
[Show abstract][Hide abstract] ABSTRACT: We report in this piece of work the synthesis of biosilica hybrid materials able to develop from hydrophobic polyesters to intermediate hydrophilic polyesters and then to polyamides through an O-N acyl transfer shift. After demonstration of the O-N acyl shift on a model depsipeptide anchored in the pores of ordered mesoporous silica (OMS), the preparation of three bio oligomer silica hybrid materials is described. The oligomerisation of protected serine lactones was initiated in the pores of carboxylate functionalized OMS. Hybrid protected serine polyester OMS were obtained and could be converted (i) into cationic polyesters, which could be switched (ii) into neutral polyamide OMS in basic conditions through a multi O-N acyl shift.
[Show abstract][Hide abstract] ABSTRACT: A novel strategy for the synthesis of peptide–bioorganic hybrid silica materials is reported. It relies on the design of hybrid peptides bearing a trialkoxysilane group. These reactive monomers are covalently attached to silica without requiring any prior surface modification. As an example, a bioorganic–inorganic OMS aldolisation catalyst is prepared using the peptide sequence Pro-Pro-Asp-Lys.
[Show abstract][Hide abstract] ABSTRACT: The use of meta-nitrobenzyl alcohol (NBA) as matrix allows to obtain better spectra than those recorded with the more popular matrix glycerol (G) in the analysis of fulvene derivatives by FAB mass spectrometry. The influence of NBA on the mass spectra corresponds to its oxidative properties. Finally, the necessary structural information was easily obtained through tandem mass spectrometry experiments.
[Show abstract][Hide abstract] ABSTRACT: The straightforward synthesis of a double-lasso macrocycle from a nonsymmetrical [c2]daisy chain, using the copper(I)-catalyzed Huisgen alkyne-azide 1,3-dipolar cycloaddition, is described. The preparation of the nonsymmetrical alkyne azide [c2]daisy chain precursor was realized in situ via the exchange of the monomers contained in both symmetrical alkyne and azide [c2]daisy chains and was followed by mass spectrometry.
[Show abstract][Hide abstract] ABSTRACT: A straightforward synthesis of comb-polypeptides of repeated peptide sequences was developed. These polypeptides were obtained by ROP of defined NCA without any post-polymerization grafting. The key to this strategy relies on the preparation of pure NCA bearing a peptide sequence on its side chain, by an original solid supported methodology.
Full-text · Article · Nov 2012 · Chemical Communications
[Show abstract][Hide abstract] ABSTRACT: We have evaluated the laser desorption ionization-mass spectrometry (LDI-MS) performance of six nanostructured silicon surfaces of different morphologies and chemical functionalizations. The substrates have been synthesized either by metal-assisted etching method or by vapor-liquid-solid (VLS) growth technique. In addition to the commercial NALDITM target plates, serving as reference, the home-made surfaces have been evaluated in mass spectrometry experiments conducted with peptide solutions mimicking tryptic digests. LDI surfaces synthesized by metal-assisted etching method were the most efficient in terms of signal intensities and number of detected peptides. The surface providing the best LDI-MS performance was composed of two nanostructured layers. Interestingly, we also observed a significant influence of the type of organic coating (hydrocarbon vs fluorocarbon) on peptide ionization discrimination.
Full-text · Article · Nov 2012 · Analytical Chemistry
[Show abstract][Hide abstract] ABSTRACT: The present study was conducted to assess the structures of the main unknown oxygenated metabolites of EAPB0203. The first step was to assign all the 1H and 13C nuclear magnetic resonances (NMR) of both EAPB0203 and its demethylated metabolite (EAPB0202) to the corresponding atoms in their molecular structures and to elucidate the fragmentation pathways for the [M+H]+ ions of these compounds using high-resolution mass spectrometry (MS). MS-MS spectra showed that both protonated molecules possessing an even number of electrons were unexpectedly losing radicals such as H•, CH3• or even C7H7• giving stable radical cations. In vitro metabolism studies were investigated in rat and dog liver microsomes and in the filamentous fungus Cunninghamella elegans. Structural elucidation of six oxygenated metabolites was performed based on (i) their fragmentation pathways in liquid chromatography-MS/MS analyses; (ii) comparison of their changes in their molecular masses and fragment ions with those of the parent drugs; and (iii) the results of on-line H/D exchange experiments which provided additional evidence in differentiating hydoxylated metabolites from N-oxides. Structures of the metabolites were elucidated by LC-MS/MS and comparison with synthetic standards; structures of these standards were confirmed using 1- and 2-D1 H NMR spectroscopy.
Full-text · Article · Oct 2012 · Analytical Chemistry
[Show abstract][Hide abstract] ABSTRACT: We report on the simple deposition of Stöber silica nanoparticles (SiO(2) NPs) on conventional MALDI target plate for high throughput laser desorption/ionization mass spectrometry (LDI-MS) analyses of peptide mixtures with sensitivity in the femtomolar range. This low-cost easily prepared material allowed straightforward LDI experiments by deposition of the studied samples directly onto a pre-spotted MALDI plate. This analytical strategy can be performed in any laboratory equipped with a MALDI-TOF instrument. All key benefits of organic matrix-free technologies were satisfied while maintaining a high level of detection performances (sensitivity and reproducibility/repeatability). In particular, sample preparation was simple and detection in the low mass range was not hampered by matrix ions. Imaging studies were undertaken to query sample dispersion into the inert SiO(2) NPs and to help into the search of the best experimental conditions producing homogeneous analyte distribution within the deposit. In contrast to commercial disposable LDI targets designed for single use and requiring an adaptor such as NALDI™, the proposed SiO(2) NPs pre-spotting on a MALDI target plate allowed very easily switching between MALDI and LDI experiments. They can be conducted either simultaneously (positions with an organic matrix or SiO(2) NPs) or in the row (support prepared in advance, stored and washed after use). The overall cost and versatility of the methodology made it very attractive to MALDI users in many domains (peptidomics, proteomics, metabolomics).
[Show abstract][Hide abstract] ABSTRACT: Sulfation of tyrosine residues is a key posttranslational modification in the regulation of various cellular processes. As such, the detection and localization of tyrosine sulfation is an essential step toward the elucidation of the physiological and pathological roles of this process. Despite substantial advances, intact sulfated peptides are still difficult to detect by MALDI-MS due to the extreme lability of the sulfo-moiety. The present report demonstrates for the first time how intact sulfated peptides can be directly and specifically detected by MALDI-MS in positive reflectron mode by using pyrenemethylguanidine (pmg) as a noncovalent derivatizing agent and an ionization enhancer. This new method allows the determination of the degree of sulfation of sulfopeptides pure or in mixtures. Moreover, the observation of specific peaks in the mass spectra enables a rapid and unambiguous discrimination between phospho- and sulfopeptides.
[Show abstract][Hide abstract] ABSTRACT: We report on the simple application of a new nanostructured silicon (NanoSi) substrate as laser desorption/ionization (LDI)-promoting surface for high-throughput identification of protein tryptic digests by a rapid MS profiling and subsequent MS/MS analysis. The NanoSi substrate is easily prepared by chemical etching of crystalline silicon in NH(4)F/HNO(3)/AgNO(3) aqueous solution. To assess the LDI performances in terms of sensitivity, repeatability and robustness, the detection of small synthetic peptides (380-1700Da) was investigated. Moreover, peptide sequencing was tackled. Various tryptic synthetic peptide mixtures were first characterized in MS and MS/MS experiments carried out on a single deposit. Having illustrated the capability to achieve peptide detection and sequencing on these ionizing surfaces in the same run, protein tryptic digests from Cytochrome C, β-Casein, BSA and Fibrinogen were then analyzed in the femtomolar range (from 50 fmol for Cytochrome C down to 2 fmol for Fibrinogen). Comparison of the NanoSi MS and MS/MS data with those obtained with sample conditioned in organic matrix demonstrated a great behavior for low mass responses. We demonstrated the capability of LDI on NanoSi to be a complementary method to MALDI peptide mass fingerprinting ensuring determination of peptide molecular weights and sequences for more efficient protein database searches.
Full-text · Article · Jan 2012 · Journal of proteomics
[Show abstract][Hide abstract] ABSTRACT: By screening a data set of 392 synthetic peptides MS/MS spectra, we found that a known C-terminal rearrangement was unexpectedly frequently occurring from monoprotonated molecular ions in both ESI and MALDI tandem mass spectrometry upon low and high energy collision activated dissociations with QqTOF and TOF/TOF mass analyzer configuration, respectively. Any residue localized at the C-terminal carboxylic acid end, even a basic one, was lost, provided that a basic amino acid such arginine and to a lesser extent histidine and lysine was present in the sequence leading to a fragment ion, usually depicted as (b(n-1) + H(2)O) ion, corresponding to a shortened non-scrambled peptide chain. Far from being an epiphenomenon, such a residue exclusion from the peptide chain C-terminal extremity gave a fragment ion that was the base peak of the MS/MS spectrum in certain cases. Within the frame of the mobile proton model, the ionizing proton being sequestered onto the basic amino acid side chain, it is known that the charge directed fragmentation mechanism involved the C-terminal carboxylic acid function forming an anhydride intermediate structure. The same mechanism was also demonstrated from cationized peptides. To confirm such assessment, we have prepared some of the peptides that displayed such C-terminal residue exclusion as a C-terminal backbone amide. As expected in this peptide amide series, the production of truncated chains was completely suppressed. Besides, multiply charged molecular ions of all peptides recorded in ESI mass spectrometry did not undergo such fragmentation validating that any mobile ionizing proton will prevent such a competitive C-terminal backbone rearrangement. Among all well-known nondirect sequence fragment ions issued from non specific loss of neutral molecules (mainly H(2)O and NH(3)) and multiple backbone amide ruptures (b-type internal ions), the described C-terminal residue exclusion is highly identifiable giving raise to a single fragment ion in the high mass range of the MS/MS spectra. The mass difference between this signal and the protonated molecular ion corresponds to the mass of the C-terminal residue. It allowed a straightforward identification of the amino acid positioned at this extremity. It must be emphasized that a neutral residue loss can be misattributed to the formation of a y(m-1) ion, i.e., to the loss of the N-terminal residue following the a(1)-y(m-1) fragmentation channel. Extreme caution must be adopted when reading the direct sequence ion on the positive ion MS/MS spectra of singly charged peptides not to mix up the attribution of the N- and C-terminal amino acids. Although such peculiar fragmentation behavior is of obvious interest for de novo peptide sequencing, it can also be exploited in proteomics, especially for studies involving digestion protocols carried out with proteolytic enzymes other than trypsin (Lys-N, Glu-C, and Asp-N) that produce arginine-containing peptides.
Full-text · Article · Nov 2011 · Journal of the American Society for Mass Spectrometry
[Show abstract][Hide abstract] ABSTRACT: The synthesis and use of novel polymer-supported reagents for disulfide bond formation is described. This family of supported reagents consists of a series of oxidized methionines grafted onto a solid support. Their cost and the simplicity of their preparation through N-carboxyanhydride polymerization on beads make them reactants of choice for the formation of disulfide bridges in peptides.
No preview · Article · Sep 2011 · Chemistry - An Asian Journal
[Show abstract][Hide abstract] ABSTRACT: AbstractWhen synthesizing arylpiperazine library modified with N-acylated amino acid derivatives (e.g., cyclized aspartic acid, cyclized glutamic acid, proline) we wished to rapidly determine
the way of cyclization of N-acylated glutamic acid derivatives. During concomitant cleavage and cyclization two alternative routes were possible—either
formation of six-member imide (glutarimide) or five-member lactam. Application of MS/MS and 1H NMR method allowed us to establish that cyclization of N-acylated glutamic acid derivatives preceded to lactams—N-acylated pyroglutamic acid derivatives.
Graphical AbstractUnder acidic treatment the Lantern bound N-acylated glutamic acid derivatives underwent concomitant cleavage and cyclization. Herein, we studied two alternative cyclization
routes—imides, lactams. It was found that cyclization of N-acylated glutamic acid derivatives yielded pyroglutamic acid derivatives (Structure C).
KeywordsSolid-phase synthesis–Aspartic acid cyclization–Glutamic acid cyclization–Pyroglutamic acid–Mass spectrometry–SynPhase Lanterns
No preview · Article · Jun 2011 · International Journal of Peptide Research and Therapeutics