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High-abundance Proteins (high-abundance + protein)
Selected AbstractsMicro-high-performance liquid chromatography platform for the depletion of high-abundance proteins and subsequent on-line concentration/capturing of medium and low-abundance proteins from serum.ELECTROPHORESIS, Issue 16 2008Application to profiling of protein expression in healthy, osteoarthritis sera by 2-D gel electrophoresis No abstract is available for this article. [source] Removal of high-abundance proteins for nuclear subproteome studies in rice (Oryza sativa) endospermELECTROPHORESIS, Issue 3 2008Guosheng Li Abstract Endosperm is a highly specialized storage organ with three sets of genomes. It is one of the most economically important organs in plants. Endosperm development involves parental imprinting and endoreduplication. A thorough study of the endosperm proteome, particularly the nuclear proteome, may provide critical insight into the regulation of seed development. Unfortunately, endosperm is extremely rich in starch grains and protein bodies of different sizes, making proteome studies on nonstorage proteins, particularly the low-abundance proteins, very challenging. Here we have developed a chromatographic method to remove large starch grains and an electrophoresis method to recover low-abundance proteins, respectively. Using these methods, we have identified 468 proteins from the nuclear enriched fraction of rice endosperm, including transcription factors, histone modification proteins, kinetochore proteins, centromere/microtubule binding proteins, and transposon proteins. Among the 468 proteins, 208 (44%) are hypothetical proteins, indicating that the endosperm proteome is poorly explored. In addition, analyses of the MS/MS data using BioWorks 3.1 have identified 59 putative acetylated proteins and 40 putative methylated proteins. Our studies have developed a method to remove starch grains and recover low-abundance proteins, respectively. The methods should be applicable to other organisms. [source] Large scale depletion of the high-abundance proteins and analysis of middle- and low-abundance proteins in human liver proteome by multidimensional liquid chromatographyPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 5 2008Mingxia Gao Abstract An unbiased method for large-scale depletion of high-abundance proteins and identification of middle- or low-abundance proteins by multidimensional LC (MDLC) was demonstrated in this paper. At the protein level, the MDLC system, coupling the first dimensional strong cation exchange (SCX) chromatography with the second dimensional RP-HPLC, instead of immunoaffinity technology, was used to deplete high-abundance proteins. Sixty-two fractions from SCX were separated further by RPLC. UV absorption spectra were observed to differentiate high-abundance proteins from middle- or low-abundance proteins. After the depletion of high-abundance proteins, middle- or low-abundance proteins were enriched, digested, and separated by online 2D-micro-SCX/cRPLC. The eluted peptides were deposited on the MALDI target and detected by MALDI-TOF/TOF MS. This depletion strategy was applied to the proteome of the normal human liver (NHL) provided by the China Human Liver Proteome Project (CHLPP). In total, 58 high-abundance proteins were depleted in one experiment. The strategy increases greatly the number of identified proteins and around 1213 proteins were identified, which was about 2.7 times as that of the nondepletion method. [source] A method for the selective isolation and enrichment of carrier protein-bound low-molecular weight proteins and peptides in the bloodPROTEOMICS - CLINICAL APPLICATIONS, Issue 2 2007Serena Camerini Abstract The low molecular weight (LMW) region of the circulatory proteome, thought to contain a rich source of biomarkers, resides in vivo, in a complexed state with larger, highly abundant resident proteins. Consequently, serum fractionation approaches that deplete the high-abundance proteins under native conditions will remove much of the LMW proteome. We describe a new strategy to systematically collect, isolate and enrich the LMW molecules that would be otherwise eliminated during the depletion of high-abundance circulatory proteins based on continuous elution electrophoresis. We employ strong denaturing conditions to disrupt association with the high-abundance carrier proteins followed by fractionation and removal of SDS. Under denaturation, the LMW molecules were effectively stripped from the highly abundant carrier proteins. We then removed the SDS by ion exchange matrix sequestration and concentrated the fractions. The outcome is a series of SDS-free fractions of LMW molecules. The isolated fractions were then analyzed by enzymatic digestion followed by LC-MS/MS analysis. The yield of multiple peptide hits as well as the total number of identifications significantly increased (50%) compared to unfractionated serum. The method yielded a 30% higher number of low-abundance serum proteins compared to direct sequencing of unfractionated serum. [source] Introducing proteomics in the undergraduate curriculum: A simple 2D gel electrophoresis exercise with serum proteinsBIOCHEMISTRY AND MOLECULAR BIOLOGY EDUCATION, Issue 1 2010Thomas D. Kim Abstract Two-dimensional gel electrophoresis (2DGE) remains an important tool in the study of biological systems by proteomics. While the use of 2DGE is commonplace in research publications, there are few instructional laboratories that address the use of 2DGE for analyzing complex protein samples. One reason for this lack is the fact that the preparation of samples for 2DGE is a complex and difficult process that can commonly yield gels of poor quality and resolution. In this experiment, we use a serum-based sample to mitigate many of the sample preparation issues that occur in cell-based sample preparations and incorporate a protein precipitation method that was developed to address the problem of high-abundance proteins and dynamic range in serum proteomics research. By focusing on 2DGE apart from many other facets of proteomic experimental design, students have the opportunity to gain fruitful experience in the use of this workhorse proteomics technique. This simplified focus also makes this exercise accessible to biochemistry instructors who are not active in proteomics; the requisite techniques may require some new equipment (i.e. an isoelectric focusing apparatus), but this exercise focuses on using familiar techniques (primarily electrophoresis) to cross the threshold of a new field, proteomics. [source] Limitation of immunoaffinity column for the removal of abundant proteins from plasma in quantitative plasma proteomicsBIOMEDICAL CHROMATOGRAPHY, Issue 5 2009Tomoko Ichibangase Abstract In plasma proteomics, before a proteome analysis, it is essential to prepare protein samples without high-abundance proteins, including albumin, via specific preparation techniques, such as immunoaffinity capture. However, our preliminary experiments suggested that functional changes with use alter the ability of the immunoaffinity column. Thus, in this study, to evaluate the changes of the removal ability of abundant proteins from plasma by the immunoaffinity column, plasma proteome analysis was performed for the long-term test for the reproducibility of the affinity column using the fluorogenic derivatization,liquid chromatography,tandem mass spectrometry method combined with an IgY column. The specific adsorption for albumin decreased with an increase in the number of the column usage before its expiration date. Moreover, it was demonstrated that hydrophobic high molecular weight compounds in plasma adsorbed onto the column materials surface contributed to the functional changes from specific immunoaffinity adsorption into hydrophobic interaction. These results suggested that, in quantitative plasma proteomics studies, it is important to keep in mind the risk of not only the nonselective loss but also the changes in the adsorption ability of the immunoafinity column. Copyright © 2008 John Wiley & Sons, Ltd. [source] |