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Peptide Ions (peptide + ion)
Selected AbstractsHigh-sensitivity analysis of specific peptides in complex samples by selected MS/MS ion monitoring and linear ion trap mass spectrometry: Application to biological studiesJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 11 2007Inmaculada Jorge Abstract Mass spectrometry (MS) is a technique of paramount importance in Proteomics, and developments in this field have been possible owing to novel MS instrumentation, experimental strategies, and bioinformatics tools. Today it is possible to identify and determine relative expression levels of thousands of proteins in a biological system by MS analysis of peptides produced by proteolytic digestion. In some situations, however, the precise characterization of a particular peptide species in a very complex peptide mixture is needed. While single-fragment ion-based scanning modes such as selected ion reaction monitoring (SIRM) or consecutive reaction monitoring (CRM) may be highly sensitive, they do not produce MS/MS information and their actual specificity must be determined in advance, a prerequisite that is not usually met in a basic research context. In such cases, the MS detector may be programmed to perform continuous MS/MS spectra on the peptide ion of interest in order to obtain structural information. This selected MS/MS ion monitoring (SMIM) mode has a number of advantages that are fully exploited by MS detectors that, like the linear ion trap, are characterized by high scanning speeds. In this work, we show some applications of this technique in the context of biological studies. These results were obtained by selecting an appropriate combination of scans according to the purpose of each one of these research scenarios. They include highly specific identification of proteins present in low amounts, characterization and relative quantification of post-translational modifications such as phosphorylation and S -nitrosylation and species-specific peptide identification. Copyright © 2007 John Wiley & Sons, Ltd. [source] Off-line liquid chromatography-MALDI by with various matrices and tandem mass spectrometry for analysis of glycated human serum albumin tryptic peptidesMOLECULAR NUTRITION & FOOD RESEARCH (FORMERLY NAHRUNG/FOOD), Issue 4 2007Annunziata Lapolla Abstract Advanced glycation end-product (AGE)/peptides, arising from in vivo digestion of glycated proteins, are biologically important compounds, due to their reactivity against circulating and tissue proteins. For information on their possible structure, in vitro glycation of HSA and its further enzymatic digestion were performed. The resulting digestion product mixture was analysed directly by MALDI MS with various matrices [2,5-dihydroxy benzoic acid (DHB) and ,-cyano-4-hydroxy cinnamic acid (CHCA)]. Alternatively, offline microbore LC prior to MALDI analysis was used, and showed that 63% of the free amino groups prone to glycation are modified, indicating the contemporary presence of unglycated peptides. This result proves that, regardless of the high glucose concentration employed for HSA incubation, glycation does not go to completion. Further studies showed that the collisionally activated decomposition of singly charged glycated peptides leads to specific fragmentation pathways, all related to the condensed glucose molecule. These unique product ions can be used as effective markers to establish the presence of a glucose molecule within a peptide ion. [source] Complementary structural information of positive- and negative-ion MSn spectra of glycopeptides with neutral and sialylated N-glycansRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 5 2006Kisaburo Deguchi Positive- and negative-ion MSn spectra of chicken egg yolk glycopeptides binding a neutral and a sialylated N-glycan were acquired by using electrospray ionization linear ion trap time-of-flight mass spectrometry (ESI-LIT-TOFMS) and collision-induced dissociation (CID) with helium as collision gas. Several characteristic differences were observed between the positive- and negative-ion CID MSn (n,=,2, 3) spectra. In the positive-ion MS2 spectra, the peptide moiety was presumably stable, but the neutral N-glycan moiety caused several B-type fragmentations and the sialylated N-glycan almost lost sialic acid(s). In contrast, in the negative-ion MS2 spectra, the peptide moiety caused several side-chain and N-glycan residue (e.g., N -acetylglucosamine (GlcNAc) residue) fragmentations in addition to backbone cleavages, but the N-glycan moieties were relatively stable. The positive-ion MS3 spectra derived from the protonated peptide ion containing a GlcNAc residue (203.1,Da) provided enough information to determine the peptide amino-acid sequence including the glycosylation site, while the negative-ion MS3 spectra derived from the deprotonated peptide containing a 0,2X1 -type cross-ring cleavage (83.1,Da) complicated the peptide sequence analysis due to side-chain and 0,2X1 residue related fragmentations. However, for the structural information of the N-glycan moiety of the glycopeptides, the negative-ion CID MS3 spectra derived from the deprotonated 2,4A6 -type cross-ring cleavage ion (neutral N-glycan) or the doubly deprotonated B6 -type fragment ion (sialylated N-glycan) are more informative than are those of the corresponding positive-ion CID MS3 spectra. Thus, the positive-ion mode of CID is useful for the analyses of peptide amino-acid sequences including the glycosylation site. The negative-ion mode of CID is especially useful for sialylated N-glycan structural analysis. Therefore, in the structural analysis of N-glycopeptides, their roles are complementary. Copyright © 2006 John Wiley & Sons, Ltd. [source] Dissociation mechanisms and implication for the presence of multiple conformations for peptide ions with arginine at the C-terminus: time-resolved photodissociation studyJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 7 2010So Hee Yoon Abstract Time-resolved photodissociation (PD) patterns of singly protonated peptides with arginine at the C-terminus (C-arg peptide ions) have been used to classify the dissociation channels into two categories, i.e. high-energy channels generating v, w and x and low-energy ones generating b, y and z. x + 1 formed by C,CO cleavage seems to be the intermediate ion in high-energy channels just as a + 1 is for N-arg peptide ions. Difference in time-resolved pattern indicates that the two sets of channels, high- and low-energy ones, are not in direct competition. Noncompetitive dissociation is also indicated by the observation of anomalous effect of matrix used in matrix-assisted laser desorption ionization, a cooler matrix generating more high-energy product ions both in spontaneous dissociation and in PD. Results from detailed investigation suggest that the two sets of channels start from two (or more) different conformations. Copyright © 2010 John Wiley & Sons, Ltd. [source] Peak capacity of ion mobility mass spectrometry: the utility of varying drift gas polarizability for the separation of tryptic peptidesJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 4 2004Brandon T. Ruotolo Abstract Ion mobility mass spectrometry (IM-MS) peptide mass mapping experiments were performed using a variety of drift gases (He, N2, Ar and CH4). The drift gases studied cover a range of polarizabilities ((0.2,2.6) × 10,24 cm3) and the peak capacities obtained for tryptic peptides in each gas are compared. Although the different gases exhibit similar peak capacities (5430 (Ar) to 7580 (N2)) in some cases separation selectivity presumably based on peptide conformers (or conformer populations), is observed. For example the drift time profiles observed for some tryptic peptide ions from aldolase (rabbit muscle) show a dependence on drift gas. The transmission of high-mass ions (m/z > 2000) is also influenced by increased scattering cross-section of the more massive drift gases. Consequently the practical peak capacity for IM-MS separation cannot be assumed to be solely a function of resolution and the ability of a gas to distribute signals in two-dimensional space; rather, peak capacity estimates must account for the transmission losses experienced for peptide ions as the drift gas mass increases. Copyright © 2004 John Wiley & Sons, Ltd. [source] A novel tandem quadrupole mass spectrometer allowing gaseous collisional activation and surface induced dissociationJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 12 2001Shabaz Mohammed Abstract A novel tandem quadrupole mass spectrometer is described that enables gaseous collision-induced dissociation (CID) and surface-induced dissociation (SID) experiments. The instrument consists of a commercially available triple quadrupole mass spectrometer connected to an SID region and an additional, orthogonal quadrupole mass analyser. The performance of the instrument was evaluated using leucine-enkephalin, allowing a comparison between CID and SID, and with previous reports of other SID instruments. The reproducibility of SID data was assessed by replicate determinations of the collision energy required for 50% dissociation of leucine-enkephalin; excellent precision was observed (standard deviation of 0.6 eV) though, unexpectedly, the reproducibility of the equivalent figure for CID was superior. Several peptides were analysed using SID in conjunction with liquid secondary-ion mass spectrometry or electrospray; a comparison of the fragmentation of singly protonated peptide ions and the further dissociation of y-type fragments was consistent with the equivalence of the latter fragments to protonated peptides. Few product ions attributable to high-energy cleavages of amino acid side-chains were observed. The SID properties were investigated of a series of peptides differing only in the derivatization of a cysteine residue; similar decomposition efficiencies were observed for all except the cysteic acid analogue, which demonstrated significantly more facile fragmentation. Copyright © 2001 John Wiley & Sons, Ltd. [source] Gas-phase basicities for ions from bradykinin and its des-arginine analoguesJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 8 2001Nigel P. Ewing Abstract Apparent gas-phase basicities (GBapps) for [M + H]+ of bradykinin, des-Arg1 -bradykinin and des-Arg9 -bradykinin have been assigned by deprotonation reactions of [M + 2H]2+ in a Fourier transform ion cyclotron resonance mass spectrometer. With a GBapp of 225.8 ± 4.2 kcal mol,1, bradykinin [M + H]+ is the most basic of the ions studied. Ions from des-Arg1 -bradykinin and des-Arg9 -bradykinin have GBapp values of 222.8 ± 4.3 kcal mol,1 and 214.9 ± 2.3 kcal mol,1, respectively. One purpose of this work was to determine a suitable reaction efficiency ,break point' for assigning GBapp values to peptide ions using the bracketing method. An efficiency value of 0.1 (i.e. approximately 10% of all collisions resulting in a deprotonation reaction) was used to assign GBapps. Support for this criterion is provided by the fact that our GBapp values for des-Arg1 -bradykinin and des-Arg9 -bradykinin are identical, within experimental error, to literature values obtained using a modified kinetic method. However, the GBapps for bradykinin ions from the two studies differ by 10.3 kcal mol,1. The reason for this is not clear, but may involve conformation differences produced by experimental conditions. The results may be influenced by salt-bridge conformers and/or by conformational changes caused by the use of a proton-bound heterodimer in the kinetic method. Factors affecting the basicities of these peptide ions are also discussed, and molecular modeling is used to provide information on protonation sites and conformations. The presence of two highly basic arginine residues on bradykinin results in its high GBapp, while the basicity of des-Arg1 -bradykinin ions is increased by the presence of two proline residues at the N-terminus. The proline residue in the second position folds the peptide chain in a manner that increases intramolecular hydrogen bonding to the protonated N-terminal amino group of the proline at the first position. Copyright © 2001 John Wiley & Sons, Ltd. [source] Assessing a novel microfluidic interface for shotgun proteome analysesJOURNAL OF SEPARATION SCIENCE, JSS, Issue 10 2007An Staes Abstract Microfluidic interfaces coupled to ESI mass spectrometers hold great potential for proteomics as they have been shown to augment the overall sensitivity of measurements and require only a minimum of operator manipulations as compared to conventional nano-LC interfaces. Here, we evaluated a new type of HPLC-Chips holding larger enrichment columns (thus an increased sample loading capacity) for gel-free proteome studies. A tryptic digest of a human T-cell proteome was fractionated by strong cation exchange chromatography and selected fractions were analyzed by MS/MS on an IT mass spectrometer using both the new HPLC-Chip as well as a conventional nano-LC-MS/MS interface. Our results indicate that the HPLC-Chip is capable of handling very complex peptide mixtures and, in fact, leads to the identification of more peptides and proteins as compared to when a conventional interface was used. The HPLC-Chip preferentially produced doubly charged tryptic peptides. We further show that MS/MS spectra of doubly charged tryptic peptide ions are more readily identified by MASCOT as compared to those from triply charged precursors and thus argue that besides the improved chromatographic conditions provided by the HPLC-Chip, its peptide charging profile might be a secondary factor leading to an increased proteome coverage. [source] Fragmentation pathways of protonated peptidesMASS SPECTROMETRY REVIEWS, Issue 4 2005Béla Paizs Abstract The fragmentation pathways of protonated peptides are reviewed in the present paper paying special attention to classification of the known fragmentation channels into a simple hierarchy defined according to the chemistry involved. It is shown that the ,mobile proton' model of peptide fragmentation can be used to understand the MS/MS spectra of protonated peptides only in a qualitative manner rationalizing differences observed for low-energy collision induced dissociation of peptide ions having or lacking a mobile proton. To overcome this limitation, a deeper understanding of the dissociation chemistry of protonated peptides is needed. To this end use of the ,pathways in competition' (PIC) model that involves a detailed energetic and kinetic characterization of the major peptide fragmentation pathways (PFPs) is proposed. The known PFPs are described in detail including all the pre-dissociation, dissociation, and post-dissociation events. It is our hope that studies to further extend PIC will lead to semi-quantative understanding of the MS/MS spectra of protonated peptides which could be used to develop refined bioinformatics algorithms for MS/MS based proteomics. Experimental and computational data on the fragmentation of protonated peptides are reevaluated from the point of view of the PIC model considering the mechanism, energetics, and kinetics of the major PFPs. Evidence proving semi-quantitative predictability of some of the ion intensity relationships (IIRs) of the MS/MS spectra of protonated peptides is presented. © 2004 Wiley Periodicals, Inc., Mass Spec Rev 24:508,548, 2005 [source] De novo sequencing of peptides by MS/MSPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 4 2010Joerg Seidler Abstract The current status of de novo sequencing of peptides by MS/MS is reviewed with focus on collision cell MS/MS spectra. The relation between peptide structure and observed fragment ion series is discussed and the exhaustive extraction of sequence information from CID spectra of protonated peptide ions is described. The partial redundancy of the extracted sequence information and a high mass accuracy are recognized as key parameters for dependable de novo sequencing by MS. In addition, the benefits of special techniques enhancing the generation of long uninterrupted fragment ion series for de novo peptide sequencing are highlighted. Among these are terminal 18O labeling, MSn of sodiated peptide ions, N-terminal derivatization, the use of special proteases, and time-delayed fragmentation. The emerging electron transfer dissociation technique and the recent progress of MALDI techniques for intact protein sequencing are covered. Finally, the integration of bioinformatic tools into peptide de novo sequencing is demonstrated. [source] A proteomic approach combining MS and bioinformatic analysis for the detection and identification of biomarkers of administration of exogenous human growth hormone in humansPROTEOMICS - CLINICAL APPLICATIONS, Issue 8 2009Joshua Boateng Abstract An integrated MS-based proteomic approach is described that combines MALDI-MS and LC-MS with artificial neural networks for the identification of protein and peptide biomarkers associated with recombinant human growth hormone (rhGH) administration. Serum from exercised males administered with rhGH or placebo was analysed using ELISA to determine insulin-like growth factor-I concentrations. Diluted serum from rhGH- and placebo-treated subjects was analysed for protein biomarkers by MALDI-MS, whereas LC-MS was used to analyse tryptically digested ACN-depleted serum extracts for peptide biomarkers. Ion intensities and m/z values were used as inputs to artificial neural networks to classify samples into rhGH- and placebo-treated groups. Six protein ions (MALDI-MS) correctly classified 96% of samples into their respective groups, with a sensitivity of 91% (20 of 22 rhGH treated) and specificity of 100% (24 of 24 controls). Six peptide ions (LC-MS) were also identified and correctly classified 93% of samples with a sensitivity of 90% (19 of 21 rhGH treated) and a specificity of 95% (20 of 21 controls). The peptide biomarker ion with the highest significance was sequenced using LC-MS/MS and database searching and found to be associated with leucine-rich ,-2-glycoprotein. [source] The nature of collision-induced dissociation processes of doubly protonated peptides: comparative study for the future use of matrix-assisted laser desorption/ionization on a hybrid quadrupole time-of-flight mass spectrometer in proteomicsRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 22 2001R. Cramer Comparative MS/MS studies of singly and doubly charged electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI) precursor peptide ions are described. The spectra from these experiments have been evaluated with particular emphasis on the data quality for subsequent data processing and protein/amino acid sequence identification. It is shown that, once peptide ions are formed by ESI or MALDI, their charge state, as well as the collision energy, is the main parameter determining the quality of collision-induced dissociation (CID) MS/MS fragmentation spectra of a given peptide. CID-MS/MS spectra of singly charged peptides obtained on a hybrid quadrupole orthogonal time-of-flight mass spectrometer resemble very closely spectra obtained by matrix-assisted laser desorption/ionization post-source decay time-of-flight mass spectrometry (MALDI-PSD-TOFMS). On the other hand, comparison of CID-MS/MS spectra of either singly or doubly charged ion species shows no dependence on whether ions have been formed by ESI or MALDI. This observation confirms that, at the time of precursor ion selection, further mass analysis is effectively decoupled from the desorption/ionization event. Since MALDI ions are predominantly formed as singly charged species and ESI ions as doubly charged, the associated difference in the spectral quality of MS/MS spectra as described here imposes direct consequences on data processing, database searching using ion fragmentation data, and de novo sequencing when ionization techniques are changed. Copyright © 2001 John Wiley & Sons, Ltd. [source] | |||||||||||||