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Siu Kwan Sze - One of the best experts on this subject based on the ideXlab platform.
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Characterization of Glutamine Deamidation by Long-Length Electrostatic Repulsion-Hydrophilic Interaction Chromatography-Tandem Mass Spectrometry (LERLIC-MS/MS) in Shotgun Proteomics
Analytical chemistry, 2016Co-Authors: Aida Serra, Xavier Gallart-palau, Juan Wei, Siu Kwan SzeAbstract:Deamidation of glutamine (Gln) residues is a spontaneous or enzymatic process with significant implications in aging and human pathology. Although some methods are available to identify the γ/α-glutamyl products of deamidation, none of these methods allows the characterization of this post-translational modification (PTM) from complex biological samples by shotgun proteomics. Here we present LERLIC-MS/MS, a chromatographic strategy that uses a long (50 cm) anion-exchange capillary column operating in the Electrostatic Repulsion-hydrophilic interaction mode (ERLIC) and coupled directly to tandem mass spectrometry (MS/MS) for proteome analysis in a single injection. Profiling of soluble extracts of brain tissues by LERLIC-MS/MS distinguished for the first time γ/α-glutamyl isomers of deamidation, encountering a 1.7 γ/α-glutamyl ratio for most Gln deamidation products. A detailed analysis of any deviation from that observed ratio allowed the identification of transglutaminase-mediated γ-glutamyl isomers as i...
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improving blood plasma glycoproteome coverage by coupling ultracentrifugation fractionation to Electrostatic Repulsion hydrophilic interaction chromatography enrichment
Journal of Proteome Research, 2015Co-Authors: Sunil S Adav, Dominique P.v. De Kleijn, Ho Hee Hwa, Siu Kwan SzeAbstract:Blood plasma is considered to be an excellent source of disease biomarkers because it contains proteins, lipids, metabolites, cell, and cell-derived extracellular vesicles from different cellular origins including diseased tissues. Most secretory and membranous proteins that can be found in plasma are glycoproteins; therefore, the plasma glycoproteome is one of the major subproteomes that is highly enriched with disease biomarkers. As a result, the glycoproteome has attracted much attention in clinical proteomic research. The modification of proteins with glycans regulates a wide range of functions in biology, but profiling plasma glycoproteins on a global scale has been hampered by the presence of low stoichiometry of glycoproteins in a complex high abundance plasma proteome background and lack of effective analytical technique. This study aims to improve plasma glycoproteome coverage using pig plasma as a model sample with a two-step strategy. The first step involves fractionation of the plasma proteins using ultracentrifugation into supernatant and pellet that is believed to contain low abundant glycoproteins. In the second step, further enrichment of glycopeptides was achieved in both fractions by adopting Electrostatic Repulsion hydrophilic interaction chromatography (ERLIC) coupled to tandem mass spectrometry (LC-MS/MS) analysis. The coverage of enriched glycoproteins in supernatant, pellet, and whole plasma sample as control was compared. Using this simple sample fractionation approach by ultracentrifugation and further ERLIC enrichment technique, sample complexity was reduced and glycoproteome coverage was significantly enhanced in supernatant and pellet fractions (by >50%) compared with whole plasma sample. This study showed that when ultracentrifugation is coupled to ERLIC glycopeptides enrichment and glycoproteome identification are significantly improved. This study demonstrates the combination of ultracentrifugation and ERLIC as a useful method for discovering plasma glycoprotein disease biomarkers.
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Simultaneous Enrichment of Plasma Soluble and Extracellular Vesicular Glycoproteins Using Prolonged Ultracentrifugation-Electrostatic Repulsion-hydrophilic Interaction Chromatography (PUC-ERLIC) Approach
Molecular & cellular proteomics : MCP, 2015Co-Authors: Esther Sok Hwee Cheow, Kae Hwan Sim, Dominique P.v. De Kleijn, Chuen Neng Lee, Vitaly Sorokin, Siu Kwan SzeAbstract:Plasma glycoproteins and extracellular vesicles represent excellent sources of disease biomarkers, but laboratory detection of these circulating structures are limited by their relatively low abundance in complex biological fluids. Although intensive research has led to the development of effective methods for the enrichment and isolation of either plasma glycoproteins or extracellular vesicles from clinical materials, at present it is not possible to enrich both structures simultaneously from individual patient sample, a method that affords the identification of biomarker combinations from both entities for the prediction of clinical outcomes will be clinically useful. We have therefore developed an enrichment method for use in mass spectrometry-based proteomic profiling that couples prolonged ultracentrifugation with Electrostatic Repulsion-hydrophilic interaction chromatography, to facilitate the recovery of both glycoproteins and extracellular vesicles from nondepleted human plasma. Following prolonged ultracentrifugation, plasma glycoproteins and extracellular vesicles were concentrated as a yellow suspension, and simultaneous analyses of low abundant secretory and vesicular glycoproteins was achieved in a single LC-MS/MS run. Using this systematic prolonged ultracentrifugation-Electrostatic Repulsion-hydrophilic interaction chromatography approach, we identified a total of 127 plasma glycoproteins at a high level of confidence (FDR ≤ 1%), including 48 glycoproteins with concentrations ranging from pg to ng/ml. The novel enrichment method we report should facilitate future human plasma-based proteome and glycoproteome that will identify novel biomarkers, or combinations of secreted and vesicle-derived biomarkers, that can be used to predict clinical outcomes in human patients.
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The Use of Electrostatic Repulsion-Hydrophilic Interaction Chromatography (ERLIC) for Proteomics Research
Mass Spectrometry Letters, 2014Co-Authors: Piliang Hao, Siu Kwan SzeAbstract:Characterization and studies of proteome are challenging because biological samples are complex, with a wide dynamic range of abundance. At present the proteins are identified by digestion into peptides, with subsequent identification of the peptides by mass spectrometry (MS). MS is a powerful technique for the purpose, but it cannot identify every peptide in such complex mixtures simultaneously. For accurate analysis and quantification it is important to separate the peptides first by chro- matography into fractions of a size that MS can handle. With these less complex fractions, the probability is increased of identi- fying peptides of low abundance that would otherwise experience ion suppression effects due to the presence of peptides of high abundance. Enrichment for peptides with certain post-translational modifications helps to increase their detection rates as well. Electrostatic Repulsion-hydrophilic interaction chromatography (ERLIC) is a mixed-mode chromatographic technique which combines the use of Electrostatic Repulsion and hydrophilic interaction. This review provides an overview of ERLIC and its var- ious proteomics applications. ERLIC has been demonstrated to have good orthogonality to reverse phase liquid chromatography (RPLC), making it useful as a first dimension in multidimensional liquid chromatography (MDLC) and fractionation of digests in general. Peptides elute in order of their isoelectric points and polarity. ERLIC has also been successfully utilized for the enrichment for phosphopeptides and glycopeptides, facilitating their identification. In addition, it is promising for the study of peptide deamidation. ERLIC performs comparably well or better than established methods for these various applications, and serves as a viable and efficient workflow alternative.
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Electrostatic Repulsion-Hydrophilic Interaction Chromatography (ERLIC) versus Strong Cation Exchange (SCX) for Fractionation of iTRAQ-Labeled Peptides
Journal of proteome research, 2011Co-Authors: Piliang Hao, Jingru Qian, Yan Ren, Siu Kwan SzeAbstract:The iTRAQ technique is popular for the comparative analysis of proteins in different complex samples. To increase the dynamic range and sensitivity of peptide identification in shotgun proteomics, SCX chromatography is generally used for the fractionation of iTRAQ-labeled peptides before LC–MS/MS analysis. However, SCX suffers from clustering of similarly charged peptides and the need to desalt fractions. In this report, SCX is compared with the alternative ERLIC method for fractionating iTRAQ-labeled peptides. The simultaneous effect of Electrostatic Repulsion and hydrophilic interaction in ERLIC results in peptide elution in order of decreasing pI and GRAVY values (increasing polarity). Volatile solvents can be used. We applied ERLIC to iTRAQ-labeled peptides from rat liver tissue, and 2745 proteins and 30 016 unique peptides were identified with high confidence from three technical replicates. This was 12.9 and 49.4% higher, respectively, than was obtained using SCX. In addition, ERLIC is appreciably b...
Cunliu Zhou - One of the best experts on this subject based on the ideXlab platform.
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l arginine l lysine improves emulsion stability of chicken sausage by increasing Electrostatic Repulsion of emulsion droplet and decreasing the interfacial tension of soybean oil water
Food Hydrocolloids, 2019Co-Authors: Xiaoxu Zhu, Cheng Ning, Cunliu ZhouAbstract:Abstract The effects of l –arginine (Arg) and l –lysine (Lys) on the emulsifying properties of the salt-soluble meat proteins (SSMPs)–soybean oil emulsion (SSOE) and soybean oil–water emulsion (SWE) were evaluated to ascertain their roles in improving the emulsion stability of sausage. The results indicated that Arg/Lys addition or increasing pH increased the emulsifying activity/stability index and the absolute ζ–potential of emulsion droplets but decreased the creaming index, apparent viscosity, and mean particle size of SSOE. Amino acid analysis showed that Arg/Lys accumulated at the interface of the emulsion droplets. Arg/Lys also decreased the surface tension of SWE at 0.002% but increased the stability of SWE at 0.1% – 0.3%. In summary, Arg/Lys improved the stability of emulsified sausage by increasing the Electrostatic Repulsion of emulsion droplets and decreasing the interfacial tension of soybean oil-water.
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l–Arginine/l–lysine improves emulsion stability of chicken sausage by increasing Electrostatic Repulsion of emulsion droplet and decreasing the interfacial tension of soybean oil-water
Food Hydrocolloids, 2019Co-Authors: Xiaoxu Zhu, Cheng Ning, Cunliu ZhouAbstract:Abstract The effects of l –arginine (Arg) and l –lysine (Lys) on the emulsifying properties of the salt-soluble meat proteins (SSMPs)–soybean oil emulsion (SSOE) and soybean oil–water emulsion (SWE) were evaluated to ascertain their roles in improving the emulsion stability of sausage. The results indicated that Arg/Lys addition or increasing pH increased the emulsifying activity/stability index and the absolute ζ–potential of emulsion droplets but decreased the creaming index, apparent viscosity, and mean particle size of SSOE. Amino acid analysis showed that Arg/Lys accumulated at the interface of the emulsion droplets. Arg/Lys also decreased the surface tension of SWE at 0.002% but increased the stability of SWE at 0.1% – 0.3%. In summary, Arg/Lys improved the stability of emulsified sausage by increasing the Electrostatic Repulsion of emulsion droplets and decreasing the interfacial tension of soybean oil-water.
Veronique Nardellorataj - One of the best experts on this subject based on the ideXlab platform.
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supramolecular big bang in a single ionic surfactant water system driven by Electrostatic Repulsion from vesicles to micelles
Langmuir, 2017Co-Authors: Loïc Leclercq, Pierre Bauduin, Veronique NardelloratajAbstract:In aqueous solution, dimethyldi-n-octylammonium chloride, [DiC8][Cl], spontaneously forms dimers at low concentrations (1–10 mM) to decrease the strength of the hydrophobic–water contact. Dimers represent ideal building blocks for the abrupt edification of vesicles at 10 mM. These vesicles are fully characterized by dynamic and static light scattering, self-diffusion nuclear magnetic resonance, and freeze-fracture transmission electron microscopy. An increase in concentration leads to Electrostatic Repulsion between vesicles that explode into small micelles at 30 mM. These transitions are detected by means of surface tension, conductivity, and solubility of hydrophobic solutes as well as by isothermal titration microcalorimetry. These unusual supramolecular transitions emerge from the surfactant chemical structure that combines two contradictory features: (i) the double-chain structure tending to form low planar aggregates with low water solubility and (ii) the relatively short chains giving high hydrophi...
Albert Sickmann - One of the best experts on this subject based on the ideXlab platform.
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Highly Sensitive Phosphoproteomics by Tailoring Solid-Phase Extraction to Electrostatic Repulsion-Hydrophilic Interaction Chromatography
Analytical chemistry, 2015Co-Authors: Stefan Loroch, René P. Zahedi, Albert SickmannAbstract:In the past decade, several strategies for comprehensive phosphoproteome analysis have been introduced. Most of them combine different phosphopeptide enrichment techniques and require starting material in the milligram range, as a consequence of their insufficient sensitivity. This limitation impairs the applicability of phosphoproteomics to a wide variety of clinical research, where sample material is highly limited. Here we introduce a highly sensitive and easy-to-establish 2D bottom-up strategy for microgram-scale phosphoproteomics, based on Electrostatic Repulsion–hydrophilic interaction chromatography (ERLIC), a simple solid-phase extraction step by strong cation exchange (SCX) or reversed phase (RP), and LC-MS analysis. With only 100 μg of tryptic digested, nonstimulated HeLa protein and 45 h of LC-MS analysis time, we identified ≥7500 nonredundant and highly confident phosphorylation sites (per replicate). We assigned all phosphorylation sites to 3013 phosphoproteins, covering the entire dynamic ra...
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Glycosylation Site Analysis of Human Platelets by Electrostatic Repulsion Hydrophilic Interaction Chromatography
Clinical Proteomics, 2008Co-Authors: Urs Lewandrowski, René P. Zahedi, Katharina Lohrig, Dirk Wolters, Albert SickmannAbstract:Glycosylations range among the most common posttranslational modifications with an estimated 50% of all proteins supposed to be glycosylated. These modifications are required for essential cellular processes including cell–cell recognition, protein structure and activity, e.g., of surface receptors, as well as subcellular localization of proteins. Beside the elucidation of the carbohydrate structures, the annotation of glycosylation sites is of primary interest as a basis for subsequent functional characterization. Although mass spectrometry is the method of choice for large-scale analysis of glycosylation sites, it requires initial enrichment of glycopeptides prior mass spectrometric detection in most cases. In this paper, we present a novel approach for glycopeptide enrichment by Electrostatic Repulsion hydrophilic interaction chromatography (ERLIC). Glycopeptides were separated from the bulk of non-modified peptides and gradually eluted from the stationary phase with potential for isoform resolution. Applied to human platelets, 125 glycosylation sites on 66 proteins were identified including major platelet glycoproteins responsible for cellular function. These sites add a major contribution to the now more than 250 glycosylation sites annotated for platelets, which enable the clinically relevant design of quantification assays for platelet glycoproteins.
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Glycosylation Site Analysis of Human Platelets by Electrostatic Repulsion Hydrophilic Interaction Chromatography
Clinical Proteomics, 2008Co-Authors: Urs Lewandrowski, René P. Zahedi, Katharina Lohrig, Dirk Wolters, Albert SickmannAbstract:Introduction Glycosylations range among the most common posttranslational modifications with an estimated 50% of all proteins supposed to be glycosylated. These modifications are required for essential cellular processes including cell–cell recognition, protein structure and activity, e.g., of surface receptors, as well as subcellular localization of proteins. Beside the elucidation of the carbohydrate structures, the annotation of glycosylation sites is of primary interest as a basis for subsequent functional characterization. Although mass spectrometry is the method of choice for large-scale analysis of glycosylation sites, it requires initial enrichment of glycopeptides prior mass spectrometric detection in most cases. Materials and Methods In this paper, we present a novel approach for glycopeptide enrichment by Electrostatic Repulsion hydrophilic interaction chromatography (ERLIC). Glycopeptides were separated from the bulk of non-modified peptides and gradually eluted from the stationary phase with potential for isoform resolution. Applied to human platelets, 125 glycosylation sites on 66 proteins were identified including major platelet glycoproteins responsible for cellular function. Conclusion These sites add a major contribution to the now more than 250 glycosylation sites annotated for platelets, which enable the clinically relevant design of quantification assays for platelet glycoproteins.
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Glycosylation Site Analysis of Human Platelets by Electrostatic Repulsion Hydrophilic Interaction Chromatography
Clinical Proteomics, 2008Co-Authors: Urs Lewandrowski, René P. Zahedi, Katharina Lohrig, Dirk Wolters, Albert SickmannAbstract:Introduction Glycosylations range among the most common posttranslational modifications with an estimated 50% of all proteins supposed to be glycosylated. These modifications are required for essential cellular processes including cell–cell recognition, protein structure and activity, e.g., of surface receptors, as well as subcellular localization of proteins. Beside the elucidation of the carbohydrate structures, the annotation of glycosylation sites is of primary interest as a basis for subsequent functional characterization. Although mass spectrometry is the method of choice for large-scale analysis of glycosylation sites, it requires initial enrichment of glycopeptides prior mass spectrometric detection in most cases. Materials and Methods In this paper, we present a novel approach for glycopeptide enrichment by Electrostatic Repulsion hydrophilic interaction chromatography (ERLIC). Glycopeptides were separated from the bulk of non-modified peptides and gradually eluted from the stationary phase with potential for isoform resolution. Applied to human platelets, 125 glycosylation sites on 66 proteins were identified including major platelet glycoproteins responsible for cellular function. Conclusion These sites add a major contribution to the now more than 250 glycosylation sites annotated for platelets, which enable the clinically relevant design of quantification assays for platelet glycoproteins.
Xiaoxu Zhu - One of the best experts on this subject based on the ideXlab platform.
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l arginine l lysine improves emulsion stability of chicken sausage by increasing Electrostatic Repulsion of emulsion droplet and decreasing the interfacial tension of soybean oil water
Food Hydrocolloids, 2019Co-Authors: Xiaoxu Zhu, Cheng Ning, Cunliu ZhouAbstract:Abstract The effects of l –arginine (Arg) and l –lysine (Lys) on the emulsifying properties of the salt-soluble meat proteins (SSMPs)–soybean oil emulsion (SSOE) and soybean oil–water emulsion (SWE) were evaluated to ascertain their roles in improving the emulsion stability of sausage. The results indicated that Arg/Lys addition or increasing pH increased the emulsifying activity/stability index and the absolute ζ–potential of emulsion droplets but decreased the creaming index, apparent viscosity, and mean particle size of SSOE. Amino acid analysis showed that Arg/Lys accumulated at the interface of the emulsion droplets. Arg/Lys also decreased the surface tension of SWE at 0.002% but increased the stability of SWE at 0.1% – 0.3%. In summary, Arg/Lys improved the stability of emulsified sausage by increasing the Electrostatic Repulsion of emulsion droplets and decreasing the interfacial tension of soybean oil-water.
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l–Arginine/l–lysine improves emulsion stability of chicken sausage by increasing Electrostatic Repulsion of emulsion droplet and decreasing the interfacial tension of soybean oil-water
Food Hydrocolloids, 2019Co-Authors: Xiaoxu Zhu, Cheng Ning, Cunliu ZhouAbstract:Abstract The effects of l –arginine (Arg) and l –lysine (Lys) on the emulsifying properties of the salt-soluble meat proteins (SSMPs)–soybean oil emulsion (SSOE) and soybean oil–water emulsion (SWE) were evaluated to ascertain their roles in improving the emulsion stability of sausage. The results indicated that Arg/Lys addition or increasing pH increased the emulsifying activity/stability index and the absolute ζ–potential of emulsion droplets but decreased the creaming index, apparent viscosity, and mean particle size of SSOE. Amino acid analysis showed that Arg/Lys accumulated at the interface of the emulsion droplets. Arg/Lys also decreased the surface tension of SWE at 0.002% but increased the stability of SWE at 0.1% – 0.3%. In summary, Arg/Lys improved the stability of emulsified sausage by increasing the Electrostatic Repulsion of emulsion droplets and decreasing the interfacial tension of soybean oil-water.