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Comprehensive analysis of protein glycation is important for better understanding of its formation mechanism and biological significance. The current preconcentration methods of glycated proteome mainly depend on the reversible combination of boronic acid and cis -dihydroxy group by pH adjustment, but it has inherent limitations (e.g., poor specificity and time-consuming). Herein, for the first time, a novel enrichment method for glycated peptides is proposed based on the reversible chemical reaction between aldehyde and 1,2-aminothiol groups, in which oxidized glycated peptides are captured onto the magnetic nanoparticles via thiazolidine chemistry and then released by palladium-mediated cleavage. The method is rapid, with excellent selectivity (even at a 1:1000 molar ratio of glycated peptides/nonglycated peptides) and high sensitivity (1 fmol/μL). As a good evidence, 1549 glycated peptides were identified from glycated human serum with 94.6% specificity, providing a powerful technique for high-throughput analysis of glycated peptides.

The liver proteome can serve as a reference to better understand both disease mechanisms and possible therapeutics, since the liver is an important organ in the body that performs a large number of tasks. Here we identify the organelle proteome of C57BL/6J mouse liver nuclei as a promising strategy to enrich low abundance proteins, in the sense that analysis of whole liver cells is rather complex for current techniques and may not be suitable for proteins with low abundance. Evaluation of nucleus integrity and purity was performed to demonstrate the effectiveness of the optimized isolation procedure. The extracted nuclear proteins were identified by 2-DE MS analyses, and a total of 748 proteins were identified. Bioinformatic analyses were performed to demonstrate the physicochemical properties, cellular locations and functions of the proteins.

Hepatitis B can progress into hepatocellular carcinoma. Body irons may interfere with the clearance of hepatitis B virus (HBV) and contribute to genesis of tumor. To investigate the role of iron played in HBV-related pathogenesis, here we studied the effect of iron with different concentrations and valence states on growth of HepG2.2.15 cells and secretion of virus proteins. A strong tolerance of HepG2.2.15 cells to iron challenge was found. The concentration of hepatitis B surface antigen in cell culture medium was decreased after iron stimulation. Lower concentrations of iron facilitated hepatitis B e-antigen (HBeAg) secretion. Fe(2+) appeared more effective on HBeAg secretion than Fe(3+) did. In parallel, the differential protein profiles in HepG2.2.15 cells were studied by iTRAQ and LC-MS/MS. The differentially expressed proteins were mainly involved in stress response, signal transduction, apoptosis, etc. Four proteins (14-3-3 β/α, VCP, migration inhibitory factor, and Nup153) were verified by Western-blotting and found to be consistent with the iTRAQ data. Interestingly, nuclear import of Nuclear factor kappa B (NFκB) and its activity were found to be affected by the decreased Nup153 in iron stimulated HepG2.2.15 cells. The results may indicate possible molecular mechanism how the synergism of HBV and iron stimulation damages host liver cells., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Myocardial proteasomes are comprised of 20S core particles and 19S regulatory particles, which together carry out targeted degradation of cardiac proteins. The 19S complex is unique among the regulators of proteasomes in that it affects both the capacity and specificity of protein degradation. However, a comprehensive molecular characterization of cardiac 19S complexes is lacking. In this investigation, we tailored a multidimensional chromatography-based purification strategy to isolate structurally intact and functionally viable 19S complexes from murine hearts. Two distinct subpopulations of 19S complexes were isolated based upon (1) potency of activating 20S proteolytic activity, and (2) molecular composition using a combination of immuno-detection, two-dimensional-differential gel electrophoresis, and MS-based approaches. Heat shock protein 90 (Hsp90) was identified to be characteristic to 19S subpopulation I. The physical interaction of Hsp90 with 19S complexes was demonstrated via multiple approaches. Inhibition of Hsp90 activity using geldanamycin or BIIB021 potentiated the ability of subpopulation I to activate 20S proteasomes in the murine heart, thus demonstrating functional specificity of Hsp90 in subpopulation I. This investigation has advanced our understanding of the molecular heterogeneity of cardiac proteasomes by identifying molecularly and functionally distinct cardiac 19S complexes. The preferential association of Hsp90 with 19S subpopulation I unveils novel targets for designing proteasome-based therapeutic interventions for combating cardiac disease.

Chronic infection by hepatitis C virus (HCV) is closely correlated with serious liver diseases. Although considerable progress has been made during recent years, the mechanism of replication and pathogenesis of HCV infection are still elusive. We have applied proteomic techniques in this work to globally analyze the protein expression profiles of a human liver cell lines Huh7 in absence and presence of HCV replication, aiming at elucidating the components of HCV replication and the cellular responses to HCV replication. The protein mixtures of three subcellular fractions from Huh7 and Huh7-HCV were separated by 2-DE under various pH gradients. Differentially expressed spots were identified by MALDI-TOF MS, followed by database searching. A total of 179 comparative proteins were identified unambiguously, including proteins associated with host cytoskeleton, intracellular traffic, oxidative and ER stress, proteasome degradation, translation, apoptosis, proliferation, etc. Host proteins known to interact with HCV proteins, such as HSP27, alpha-actinin, nucleolin and eukaryotic initiation factor 4A-I, were elevated in Huh7-HCV cells. Our study provides the global information of proteomic alteration of Huh7 cells in the presence of HCV replication and the clues for further understanding of the mechanism of HCV replication and pathogenesis.

We report for the first time an expressed proteome for human hepatocellular carcinoma (HCC) in nude mice model. Most cases of human liver cancer are HCC with highly metastatic ability. Therefore, the early prediction or diagnosis and effective treatment are the key points of research. We have previously successfully established a human HCC nude mice model (LCI-D20) with high metastasis potential. To understand better the tumor biology of HCC it is worth to explore the relativity of all expressed protein profiles in the LCI-D20 HCC nude mice model. With advanced proteomics technologies, we have carried out a proteomic analysis with following stages: protein sample preparation of cancer tissue, including total cellular extraction and sequential fractionation, 2-DE and 2-D LC separation, ESI/MALDI-MS/MS identification, as well as data-dependent bioinformatics. The identified proteins were classified bioinformatically respective to their function, biological process and intracellular localization. Some important proteins found in HCC, e.g. metabolism enzymes, proteins regulating cell motility, signaling proteins, and heat shock proteins, are discussed in terms of their metastasis.

Protein post‐translational modifications (PTMs) are chemical modifications on proteins. PTMs play a key role in many cellular processes by influencing the structure of proteins and dynamically regulating their functions. Therefore, characterizing PTMs at proteome level is critical to provide invaluable insight into the functions of proteins underlying different biological processes. Advances in modern proteomics technologies including sample preparation, chromatography separation as well as mass spectrometry have propelled the PTMs proteome to further depths. During the past decade, to better examine the PTMs with high sensitivity and selectivity, our group have developed a series of MS‐based novel analytical approaches to study the protein PTMs. Herein, we mainly introduce these approaches developed by our group and discuss how to overcome the technical obstacles of studying various protein PTMs with mass spectrometry. What is the most favorite and original method developed in your research group? The qualitative and quantitative approaches developed for analysis of PTM proteome. How do you get into this specific field? Could you please share some experiences with our readers? I entered Fudan University in 2003, when the sequencing of human genome was just completed. At that time, the life science entered the post‐genomic era, the era of proteomics. My post‐doctoral work is mainly based on mass spectrometry analysis, which is the core tool for proteome research. Therefore, I began to develop mass spectrometry‐based method for proteome research since then. What is the most important personality for scientific research? Creative, persistence, and collaboration. What's your hobbies? Playing badminton. [ABSTRACT FROM AUTHOR]