Home About us Contact
|
Home About us Contact | ||
|
|
||
MALDI Mass Spectrometry (maldi + mass_spectrometry)
Selected AbstractsAmino Acids Analysis by MALDI Mass Spectrometry Using Carbon Nanotube as MatrixCHINESE JOURNAL OF CHEMISTRY, Issue 2 2005Zhang Jing Abstract Twenty common amino acids have been analyzed successfully by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) using carbon nanotubes as matrix. From the spectra, little or no background interference or fragmentation of the analytes has been observed. This method was also applied to the analysis of amino acid mixture successfully. Carbon nanotubes have some features such as large surface area to disperse the analyte molecules sufficiently and prevent the sample aggregation and strong ultraviolet absorption to transfer energy easily to the analyte molecules. The present method has potential application for the rapid and sensitive analysis of amino acids and their mixture [source] Diamond-like carbon coated polymer-based targets in microscope slide format for MALDI mass spectrometryJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 5 2010Wolfgang Winkler First page of article [source] Molecular mass determination of plasma-derived glycoproteins by ultraviolet matrix-assisted laser desorption/ionization time-of-flight mass spectrometry with internal calibrationJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 11 2002Omar Belgacem Abstract Human plasma-derived antithrombin III (AT-III), factor IX (FIX) and vitronectin (VN) were characterized as native glycoproteins and in their de- N -glycosylated form by means of MALDI mass spectrometry. The average molecular masses of the three complex glycoproteins were determined applying internal calibration with high-mass, well-defined protein calibrants. Internal calibration generated for the 47 kDa yeast protein enolase a mass precision in the continuous and delayed extraction mode of ±0.12 and ±0.022%, respectively. The achievable mass accuracy for such a high-mass, unmodified protein was in the range of 0.02% in the continuous mode, which turned out to be better than in the delayed extraction mode. Purification of all (glyco) proteins (even the calibration proteins) by means of ZipTip® technology and direct elution with a solvent system containing the appropriate MALDI matrix turned out to be a prerequisite to measure the exact molecular masses with an internal calibration. The average molecular masses of the two different forms of AT-III, namely AT-III, and AT-III,, were shown to be 57.26 and 55.04 kDa, respectively. The 2.22 kDa mass difference is attributed to the known difference in carbohydrate content at one specific site (Asn-135). After exhaustive de- N -glycosylation (by means of PNGase F) of the ,- and ,-form and subsequent MALDI-MS analysis, average molecular masses of 48.96 and 48.97 kDa, respectively, were obtained. These values are in good agreement (,0.15%) with the calculated molecular mass (49.039 kDa) of the protein part based on SwissProt data. The molecular mass of the heavily post-translational modified glycoprotein FIX was found to be 53.75 kDa with a peak width at 10% peak height of 4.5 kDa, because of the presence of many different posttranslational modifications (N - and O -glycosylation at multiple sites, sulfation, phosphorylation, hydroxylation and numerous ,-carboxyglutamic acids). MALDI-MS molecular mass determination of the native, size-exclusion chromatography-purified, VN sample revealed that the glycoprotein was present as dimer with molecular mass of 117.74 kDa, which could be corroborated by non-reducing SDS-PAGE. After sample treatment with guanidine hydrochloride and mass spectrometric analysis, a single, new main component was detected. The molecular mass turned out to be 59.45 kDa, representing the monomeric form of VN, known as V75. The determined molecular mass value was shown to be on one hand lower than from SDS-PAGE and on the other higher than the calculated amino acid sequence molecular mass (52 277 Da), pointing to the well-known SDS-PAGE bias and to considerable post-translational modifications. Further treatment of the sample with a reducing agent and subsequent MALDI-MS revealed two new components with molecular masses of 49.85 and 9.41 kDa, corresponding to V65 and V10 subunits of VN. PNGase F digest of the V75 and V65 units and MS analysis, exhibiting a molecular mass reduction of 6.37 kDa in both cases, verified the presence of a considerable amount of N -glycans. Copyright © 2002 John Wiley & Sons, Ltd. [source] Analysis of carbohydrates and glycoconjugates by matrix-assisted laser desorption/ionization mass spectrometry: An update covering the period 2001,2002MASS SPECTROMETRY REVIEWS, Issue 2 2008David J. Harvey Abstract This review is the second update of the original review on the application of MALDI mass spectrometry to the analysis of carbohydrates and glycoconjugates that was published in 1999. It covers fundamental aspects of the technique as applied to carbohydrates, fragmentation of carbohydrates, studies of specific carbohydrate types such as those from plant cell walls and those attached to proteins and lipids, studies of glycosyl-transferases and glycosidases, and studies where MALDI has been used to monitor products of chemical synthesis. Use of the technique shows a steady annual increase at the expense of older techniques such as FAB. There is an increasing emphasis on its use for examination of biological systems rather than on studies of fundamental aspects and method development and this is reflected by much of the work on applications appearing in tabular form. © 2008 Wiley Periodicals, Inc., Mass Spec Rev 27:125,201, 2008 [source] Analysis of carbohydrates and glycoconjugates by matrix-assisted laser desorption/ionization mass spectrometry: An update covering the period 1999,2000MASS SPECTROMETRY REVIEWS, Issue 4 2006David J. Harvey Abstract This review describes the use of matrix-assisted laser desorption/ionization (MALDI) mass spectrometry for the analysis of carbohydrates and glycoconjugates and continues coverage of the field from the previous review published in 1999 (D. J. Harvey, Matrix-assisted laser desorption/ionization mass spectrometry of carbohydrates, 1999, Mass Spectrom Rev, 18:349,451) for the period 1999,2000. As MALDI mass spectrometry is acquiring the status of a mature technique in this field, there has been a greater emphasis on applications rather than to method development as opposed to the previous review. The present review covers applications to plant-derived carbohydrates, N- and O- linked glycans from glycoproteins, glycated proteins, mucins, glycosaminoglycans, bacterial glycolipids, glycosphingolipids, glycoglycerolipids and related compounds, and glycosides. Applications of MALDI mass spectrometry to the study of enzymes acting on carbohydrates (glycosyltransferases and glycosidases) and to the synthesis of carbohydrates, are also covered. © 2006 Wiley Periodicals, Inc., Mass Spec Rev 25:595,662, 2006 [source] Rapid mass spectrometric analysis of 15N-Leu incorporation fidelity during preparation of specifically labeled NMR samplesPROTEIN SCIENCE, Issue 9 2008Stephanie M.E. Truhlar Abstract Advances in NMR spectroscopy have enabled the study of larger proteins that typically have significant overlap in their spectra. Specific 15N-amino acid incorporation is a powerful tool for reducing spectral overlap and attaining reliable sequential assignments. However, scrambling of the label during protein expression is a common problem. We describe a rapid method to evaluate the fidelity of specific 15N-amino acid incorporation. The selectively labeled protein is proteolyzed, and the resulting peptides are analyzed using MALDI mass spectrometry. The 15N incorporation is determined by analyzing the isotopic abundance of the peptides in the mass spectra using the program DEX. This analysis determined that expression with a 10-fold excess of unlabeled amino acids relative to the 15N-amino acid prevents the scrambling of the 15N label that is observed when equimolar amounts are used. MALDI TOF-TOF MS/MS data provide additional information that shows where the "extra" 15N labels are incorporated, which can be useful in confirming ambiguous assignments. The described procedure provides a rapid technique to monitor the fidelity of selective labeling that does not require a lot of protein. These advantages make it an ideal way of determining optimal expression conditions for selectively labeled NMR samples. [source] Epitope mapping of a monoclonal antibody against human thrombin by H/D-exchange mass spectrometry reveals selection of a diverse sequence in a highly conserved proteinPROTEIN SCIENCE, Issue 6 2002Abel Baerga-Ortiz Abstract The epitope of a monoclonal antibody raised against human thrombin has been determined by hydrogen/deuterium exchange coupled to MALDI mass spectrometry. The antibody epitope was identified as the surface of thrombin that retained deuterium in the presence of the monoclonal antibody compared to control experiments in its absence. Covalent attachment of the antibody to protein G beads and efficient elution of the antigen after deuterium exchange afforded the analysis of all possible epitopes in a single MALDI mass spectrum. The epitope, which was discontinuous, consisting of two peptides close to anion-binding exosite I, was readily identified. The epitope overlapped with, but was not identical to, the thrombomodulin binding site, consistent with inhibition studies. The antibody bound specifically to human thrombin and not to murine or bovine thrombin, although these proteins share 86% identity with the human protein. Interestingly, the epitope turned out to be the more structured of two surface regions in which higher sequence variation between the three species is seen. [source] Matrix-assisted laser desorption/ionization detection of polymerase chain reaction products by utilizing the 5,-3, exonuclease activity of Thermus aquaticus DNA polymerase,RAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 6 2003N. R. Isola The 5,-3, exonuclease activity of DNA polymerase was utilized in the polymerase chain reaction system to generate a specific signal concomitant with amplification. These signals were detected by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). This method obviates the need to perform extensive DNA purification of reaction products that is often necessary for detecting larger DNA molecules by mass spectrometry. Oligonucleotides complementary to the internal region of the amplicon are degraded by the 5,-3, exonuclease activity and the degradation products are analyzed by MALDI mass spectrometry. We refer to this assay as the Exo-taq assay or probe degradation assay. This method should be amenable to automation. Copyright © 2003 John Wiley & Sons, Ltd. [source] Cell Cycle Arrest and Apoptosis Induction by an Anticancer Chalcone EpoxideARCHIV DER PHARMAZIE, Issue 8 2010Haiyong Han Abstract Safe and effective chemotherapeutic agents for the treatment of pancreatic cancer remain elusive. We found that chalcone epoxides (1,3-diaryl-2,3-epoxypropanones) inhibited growth in two pancreatic cancer cell lines, BxPC-3 and MIA PaCa-2. Three compounds were active, with GI50 values of 5.6 to 15.8,µM. Compound 4a, 1,3-bis-(3,4,5-trimethoxyphenyl)-2,3-epoxypropanone, had an average GI50 of 14.1,µM in the NCI 60-cell-line panel. To investigate the mode of action, cell cycle analyses of BxPC-3 cells were carried out. Treatment of cells with 50,µM 4a resulted in dramatic accumulation at G2/M (61% after 12,h for 4avs. 15% for untreated cells). The cells rapidly entered apoptosis. After 12,h, 26% of cells treated with 50,µM 4a had entered apoptosis vs. 4% for cells treated with 100,µM etoposide and 2% for untreated cells. Compound 4a interfered with paclitaxel enhancement of tubulin polymerization, suggesting microtubules as the site of action. Reaction of thiol nucleophiles with 4a under basic conditions resulted in epoxide ring-opening and retroaldol fragmentation, yielding alkylated thiol. MALDI mass spectrometry showed that retroaldol reaction occurred upon treatment of ,-tubulin with 4a. The site of alkylation was identified as Cys354. Chalcone epoxides warrant further study as potential agents for treatment of cancer. [source] Control of the Molecular Weight Distribution of Petroleum Pitches via Dense-Gas ExtractionCHEMICAL ENGINEERING & TECHNOLOGY (CET), Issue 6 2007E. G. Cervo Abstract Dense-gas extraction (DGE) was used to fractionate an isotropic petroleum pitch (number-average molecular weight Mn,=,516) into oligomeric cuts. A countercurrent-flow packed column was used to effect the separation, with supercritical toluene being used as the dense-gas solvent and commercially available M-50 or A-240 pitch being used as the feed. Isothermal operation at 330, 350, and 380,°C was investigated, as well as operation with a linear positive temperature gradient (+,T), with the bottom of the column at 330 and the top at 380,°C. For isothermal operation, the molecular weight distribution of the bottom products consisted primarily of dimer (Mn,=,508) and trimer (Mn,=,759) species, with pressure changes of as little as 5 bar producing significant changes in their relative distribution, as observed by MALDI mass spectrometry. On the other hand, by operating with a +,T, we could produce a bottom product consisting primarily of trimers and tetramers (Mn,=,997). [source] Synthesis and Characterization of 2-Mono- and 1,2-Diaminocarba- closo -dodecaborates M[1-R-2-H2N- closo -CB11H10] (R=H, Ph, H2N, CyHN),CHEMISTRY - A EUROPEAN JOURNAL, Issue 4 2009Maik Finze Dr. Abstract The first primary 2-aminocarba- closo -dodecaborates [1-R-2-H2N- closo -CB11H10], (R=H (1), Ph (2)) have been synthesized by insertion reactions of (Me3Si)2NBCl2 into the trianions [7-R-7- nido -CB10H10]3,. The difunctionalized species [1,2-(H2N)2 - closo -CB11H10] (3) and 1-CyHN-2-H3N- closo -CB11H10 (H- 4) have been prepared analogously from (Me3Si)2NBCl2 and 7-H3N-7- nido -CB10H12. In addition, the preparation of [Et4N][1-H2N-2-Ph- closo -CB11H10] ([Et4N]- 5) starting from PhBCl2 and 7-H3N-7- nido -CB10H12 is described. Methylation of the [1-Ph-2-H2N- closo -CB11H10], ion (2) to produce 1-Ph-2-Me3N- closo -CB11H10 (6) is reported. The crystal structures of [Et4N]- 2, [Et4N]- 5, and 6 were determined and the geometric parameters were compared to theoretical values derived from DFT and ab initio calculations. All new compounds were studied by NMR, IR, and Raman spectroscopy, MALDI mass spectrometry, and elemental analysis. The discussion of the experimental NMR chemical shifts and of selected vibrational band positions is supported by theoretical data. The thermal properties were investigated by differential scanning calorimetry (DSC). The pKa values of 2-H3N- closo -CB11H11 (H- 1), 1-H3N- closo -CB11H10 (H- 7), and 1,2-(H3N)2 - closo -CB11H10 (H2 - 3) were determined by potentiometric titration and by NMR studies. The experimental results are compared to theoretical data (DFT and ab initio). The basicities of the aminocarba- closo -dodecaborates agree well with the spectroscopic and structural properties. [source] | ||||||||||||||||||||||