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![Profiles of chromatographic peaks. The chromatograms of M, M + 1, and M + 2 in heavy- (left) and light-labeled (right) samples are depicted in blue, purple, and brown, respectively. (A–C) Examples of outliners in the unanimous selection. (A) Case of a peak with a low signal intensity misidentified as a true peak. Sequence, K[+ 25]SAPATGGVK[+ 28]K[28 +]PHR; charge = + 4, (B) Case of an incorrect integral of interval. Sequence, DGK[+ 28]YHSIK[+ 28]EVATSVQLTLR; charge = + 3, (C) Case of differing shapes among isotopes. Sequence, LK[+ 28]QLAAEQGK[+ 28]DIR; charge = + 4.](publication/356769885/figure/fig4/AS:1097162293284887@1638595283303/Profiles-of-chromatographic-peaks-The-chromatograms-of-M-M-1-and-M-2-in-heavy-left.png)
Profiles of chromatographic peaks. The chromatograms of M, M + 1, and M + 2 in heavy- (left) and light-labeled (right) samples are depicted in blue, purple, and brown, respectively. (A–C) Examples of outliners in the unanimous selection. (A) Case of a peak with a low signal intensity misidentified as a true peak. Sequence, K[+ 25]SAPATGGVK[+ 28]K[28 +]PHR; charge = + 4, (B) Case of an incorrect integral of interval. Sequence, DGK[+ 28]YHSIK[+ 28]EVATSVQLTLR; charge = + 3, (C) Case of differing shapes among isotopes. Sequence, LK[+ 28]QLAAEQGK[+ 28]DIR; charge = + 4.
Source publication
Recent mass spectrometry (MS)-based techniques enable deep proteome coverage with relative quantitative analysis, resulting in increased identification of very weak signals accompanied by increased data size of liquid chromatography (LC)–MS/MS spectra. However, the identification of weak signals using an assignment strategy with poorer performance...
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Targeted mass spectrometry is a powerful technique for quantifying specific proteins or metabolites in complex biological samples. Accurate peak picking is a critical step as it determines the absolute abundance of each analyte by integrating the area under the picked peaks. Although automated software exists for handling such complex tasks, manual...
Citations
... Because a protein does not remain fixed in a unique structure in solution but exists as a conformational ensemble [18,19], such inconsistencies can probably be traced to no suitable protein conformation representing the average structure in solution deposited in the PDB. In this study, therefore, covalent protein labeling was used to evaluate the solvent-accessible surface of human serum albumin (HSA), a protein with numerous conformations deposited in the PDB that is commonly analyzed in pharmacokinetic studies of drug absorption, distribution, metabolism, and excretion. ...
... LC-MS/MS analyses were performed using a quadrupole Orbitrap benchtop mass spectrometer Q-Exactive (Thermo Fisher Scientific) equipped with an EASY-nLC 1000 system (Thermo Fisher Scientific) as described elsewhere [18]. Before injection, 0.1 pmol of a synthetic peptide, DRVYHIPFHL, was added as an internal standard and mixed with the same amount of tryptic peptides, and then the mixture was analyzed. ...
... Before injection, 0.1 pmol of a synthetic peptide, DRVYHIPFHL, was added as an internal standard and mixed with the same amount of tryptic peptides, and then the mixture was analyzed. MS parameters were used described previously [18] with slight modifications. Automatic gain control target values for MS1 spectra and fragment spectra were 1 Â 10 6 and 1 Â 10 5 , respectively. ...
Structural proteomics techniques are useful for identifying the binding sites of proteins. The surface of a target protein with and without a bound binding partner is artificially labeled using a hydroxy radical, deuterium, or a low-molecular-weight chemical, and the difference in the label strength with and without the bound partner is determined. Label strength maps are then prepared on the Protein Data Bank (PDB) structure to identify the binding surface. However, the surface-accessible sites determined using such structural proteomics methods are frequently inconsistent with those calculated based on PDB structures, speculating that the measurement determines chemical accessibility rather than solvent accessibility. In this study, the solvent-accessible surface of human serum albumin was analyzed using covalent protein labeling with varying concentrations of CH2O and then compared to surfaces derived from 27 PDB structures. The results indicated that inconsistencies in solvent-accessible surface area values calculated from PDB structures are not caused by the limited capabilities of liquid chromatography–mass spectrometry coupled with covalent protein painting but instead are due to the lack of PDB data representing the structures in solution.
