Cancer Serum PTM Profiling - Quantitative profiling of post-translational modifications by immunoaffinity enrichment and LC-MS/MS in cancer serum without immunodepletion
Ontology highlight
ABSTRACT:
INSTRUMENT(S): LTQ Orbitrap Velos, LTQ Orbitrap Elite
ORGANISM(S): Homo Sapiens (human)
TISSUE(S): Blood Serum
DISEASE(S): Acute Leukemia,Breast Cancer,Non-small Cell Lung Carcinoma
SUBMITTER: 
HONGBO GU
LAB HEAD: Hongbo Gu
PROVIDER: PXD002932 | Pride | 2015-12-07
REPOSITORIES: Pride
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| 012015_23168_HG_v.RAW | Raw | |||
| 012015_23169_HG_v.RAW | Raw | |||
| 012015_23170_HG_v.RAW | Raw | |||
| 012215_23171_HG_v.RAW | Raw | |||
| 012215_23172_HG_v.RAW | Raw |
Items per page: 5 1 - 5 of 138 |
Publications
Quantitative Profiling of Post-translational Modifications by Immunoaffinity Enrichment and LC-MS/MS in Cancer Serum without Immunodepletion.
Gu Hongbo H Ren Jian Min JM Jia Xiaoying X Levy Tyler T Rikova Klarisa K Yang Vicky V Lee Kimberly A KA Stokes Matthew P MP Silva Jeffrey C JC
Molecular & cellular proteomics : MCP 20151203 2
A robust method was developed and optimized for enrichment and quantitative analysis of posttranslational modifications (PTMs) in serum/plasma samples by combining immunoaffinity purification and LC-MS/MS without depletion of abundant proteins. The method was used to survey serum samples of patients with acute myeloid leukemia (AML), breast cancer (BC), and nonsmall cell lung cancer (NSCLC). Peptides were identified from serum samples containing phosphorylation, acetylation, lysine methylation, ...[more]
Similar Datasets
Project description:A robust method was developed and optimized for enrichment and quantitative analysis of posttranslational modifications (PTMs) in serum/plasma samples by combining immunoaffinity purification and LC-MS/MS without depletion of abundant proteins. The method was used to survey serum samples of patients with acute myeloid leukemia (AML), breast cancer (BC), and nonsmall cell lung cancer (NSCLC). Peptides were identified from serum samples containing phosphorylation, acetylation, lysine methylation, and arginine methylation. Of the PTMs identified, lysine acetylation (AcK) and arginine mono-methylation (Rme) were more prevalent than other PTMs. Label-free quantitative analysis of AcK and Rme peptides was performed for sera from AML, BC, and NSCLC patients. Several AcK and Rme sites showed distinct abundance distribution patterns across the three cancer types. The identification and quantification of posttranslationally modified peptides in serum samples reported here can be used for patient profiling and biomarker discovery research.
| S-EPMC4739682 | biostudies-literature
Project description:Post-translational modifications (PTMs) play a key role in numerous cellular processes by directly affecting structure, dynamics and interaction networks of target proteins. Despite their importance, our understanding of protein PTMs at the atomistic level is still largely incomplete. Molecular dynamics (MD) simulations, which provide high-resolution insight into biomolecular function and underlying mechanisms, are in principle ideally suited to tackle this problem. However, because of the challenges associated with the development of novel MD parameters and a general lack of suitable computational tools for incorporating PTMs in target protein structures, MD simulations of post-translationally modified proteins have historically lagged significantly behind the studies of unmodified proteins. Here, we present Vienna-PTM web server (http://vienna-ptm.univie.ac.at), a platform for automated introduction of PTMs of choice to protein 3D structures (PDB files) in a user-friendly visual environment. With 256 different enzymatic and non-enzymatic PTMs available, the server performs geometrically realistic introduction of modifications at sites of interests, as well as subsequent energy minimization. Finally, the server makes available force field parameters and input files needed to run MD simulations of modified proteins within the framework of the widely used GROMOS 54A7 and 45A3 force fields and GROMACS simulation package.
| S-EPMC3692090 | biostudies-literature
Project description:The identification of protein post-translational modifications (PTMs) is an increasingly important component of proteomics and biomarker discovery, but very few tools exist for performing fast and easy characterization of global PTM changes and differential comparison of PTMs across groups of data obtained from liquid chromatography-tandem mass spectrometry experiments. STRAP PTM (Software Tool for Rapid Annotation of Proteins: Post-Translational Modification edition) is a program that was developed to facilitate the characterization of PTMs using spectral counting and a novel scoring algorithm to accelerate the identification of differential PTMs from complex data sets. The software facilitates multi-sample comparison by collating, scoring, and ranking PTMs and by summarizing data visually. The freely available software (beta release) installs on a PC and processes data in protXML format obtained from files parsed through the Trans-Proteomic Pipeline. The easy-to-use interface allows examination of results at protein, peptide, and PTM levels, and the overall design offers tremendous flexibility that provides proteomics insight beyond simple assignment and counting.
| S-EPMC4240648 | biostudies-other
Project description:Gaining insight into normal cellular signaling and disease biology is a critical goal of proteomic analyses. The ability to perform these studies successfully to extract the maximum value and discovery of biologically relevant candidate biomarkers is therefore of primary importance. Many successful studies in the past have focused on total proteome analysis (changes at the protein level) combined with phosphorylation analysis by metal affinity enrichment (changes at the PTM level). Here, we use the gastric carcinoma cell line MKN-45 treated with the c-Met inhibitor SU11274 and PKC inhibitor staurosporine to investigate the most efficient and most comprehensive strategies for both total protein and PTM analysis. Under the conditions used, total protein analysis yielded few changes in response to either compound, while analysis of phosphorylation identified thousands of sites that changed differentially between the two treatments. Both metal affinity and antibody-based enrichments were used to assess phosphopeptide changes, and the data generated by the two methods was largely complementary (non-overlapping). Label-free quantitation of peptide peak abundances was used to accurately determine fold-changes between control and treated samples. Protein interaction network analysis allowed the data to be placed in a biologically relevant context, and follow-up validation of selected findings confirmed the accuracy of the proteomic data. Together, this study provides a framework for start-to-finish proteomic analysis of any experimental system under investigation to maximize the value of the proteomic study and yield the best chance for uncovering actionable target candidates.
| S-EPMC5217380 | biostudies-literature
Project description:Histones are abundant chromatin constituents carrying numerous post-translational modifications (PTMs). Such PTMs mediate a variety of biological functions, including recruitment of enzymatic readers, writers and erasers that modulate DNA replication, transcription and repair. Individual histone molecules contain multiple coexisting PTMs, some of which exhibit crosstalk, i.e. coordinated or mutually exclusive activities. Here, we present an integrated experimental and computational systems level molecular characterization of histone PTMs and PTM crosstalk. Using wild type and engineered mouse embryonic stem cells (mESCs) knocked out in components of the Polycomb Repressive Complex 2 (PRC2, Suz12(-/-)), PRC1 (Ring1A/B(-/-)) and (Dnmt1/3a/3b(-/-)) we performed comprehensive PTM analysis of histone H3 tails (50 aa) by utilizing quantitative middle-down proteome analysis by tandem mass spectrometry. We characterized combinatorial PTM features across the four mESC lines and then applied statistical data analysis to predict crosstalk between histone H3 PTMs. We detected an overrepresentation of positive crosstalk (codependent marks) between adjacent mono-methylated and acetylated marks, and negative crosstalk (mutually exclusive marks) among most of the seven characterized di- and tri-methylated lysine residues in the H3 tails. We report novel features of PTM interplay involving hitherto poorly characterized arginine methylation and lysine methylation sites, including H3R2me, H3R8me and H3K37me. Integration of the H3 data with RNAseq data by coabundance clustering analysis of histone PTMs and histone modifying enzymes revealed correlations between PTM and enzyme levels. We conclude that middle-down proteomics is a powerful tool to determine conserved or dynamic interdependencies between histone marks, which paves the way for detailed investigations of the histone code. Histone H3 PTM data is publicly available in the CrossTalkDB repository at http://crosstalkdb.bmb.sdu.dk.
| S-EPMC4974346 | biostudies-literature
Project description:We recently reported an antibody-free targeted protein quantification strategy, termed high-pressure, high-resolution separations with intelligent selection and multiplexing (PRISM), for achieving significantly enhanced sensitivity using selected reaction monitoring (SRM) mass spectrometry. Integrating PRISM with front-end IgY14 immunoaffinity depletion, sensitive detection of targeted proteins at 50-100 pg/mL levels in human blood plasma/serum was demonstrated. However, immunoaffinity depletion is often associated with undesired losses of target proteins of interest. Herein we report further evaluation of PRISM-SRM quantification of low-abundance serum proteins without immunoaffinity depletion. Limits of quantification (LOQ) at low ng/mL levels with a median coefficient of variation (CV) of ∼12% were achieved for proteins spiked into human female serum. PRISM-SRM provided >100-fold improvement in the LOQ when compared to conventional LC-SRM measurements. PRISM-SRM was then applied to measure several low-abundance endogenous serum proteins, including prostate-specific antigen (PSA), in clinical prostate cancer patient sera. PRISM-SRM enabled confident detection of all target endogenous serum proteins except the low pg/mL-level cardiac troponin T. A correlation coefficient >0.99 was observed for PSA between the results from PRISM-SRM and immunoassays. Our results demonstrate that PRISM-SRM can successfully quantify low ng/mL proteins in human plasma or serum without depletion. We anticipate broad applications for PRISM-SRM quantification of low-abundance proteins in candidate biomarker verification and systems biology studies.
| S-EPMC3733379 | biostudies-literature
Project description:Many PTMs dysregulation is known to be the major cause of many cancers including HCV induced HCC. PTMs of hepatitis C virus (HCV) regions NS3/4A, NS5A and NS5B are crucial for proper protein functions and replication that directly affect the generation of infectious virus particles and completion of its life cycle. In this study, we have performed comprehensive analysis of PTMs within HCV non-structural proteins (NS3/4A, NS5A and NS5B) through bioinformatics analysis to examine post-translational crosstalk between phosphorylation, palmitoylation, methylation, acetylation and ubiquitination sites in selected viral proteins. Our analysis has revealed many highly putative PTMs sites that are also conserved among major genotypes conferring the importance of these sites. We have also analysed viral 3D structures in their modified and unmodified forms to address extent and signatures of structural changes upon PTM. This study provides evidence that PTMs induce significant conformational changes and make viral proteins more stable. To find the potential role of PTMs in HCV induced HCC, docking analysis between selected viral proteins and p38-MAPK has been performed which also confirms their strong association with HCV induced HCC. The major findings proposed that PTMs at specific sites of HCV viral proteins could dysregulate specific pathways that cause the development of HCC.
| S-EPMC9483894 | biostudies-literature
Project description:MotivationIdentification of post-translationally modified proteins has become one of the central issues of current proteomics. Spectral library search is a new and promising computational approach to mass spectrometry-based protein identification. However, its potential in identification of unanticipated post-translational modifications has rarely been explored. The existing spectral library search tools are designed to match the query spectrum to the reference library spectra with the same peptide mass. Thus, spectra of peptides with unanticipated modifications cannot be identified.ResultsIn this article, we present an open spectral library search tool, named pMatch. It extends the existing library search algorithms in at least three aspects to support the identification of unanticipated modifications. First, the spectra in library are optimized with the full peptide sequence information to better tolerate the peptide fragmentation pattern variations caused by some modification(s). Second, a new scoring system is devised, which uses charge-dependent mass shifts for peak matching and combines a probability-based model with the general spectral dot-product for scoring. Third, a target-decoy strategy is used for false discovery rate control. To demonstrate the effectiveness of pMatch, a library search experiment was conducted on a public dataset with over 40,000 spectra in comparison with SpectraST, the most popular library search engine. Additional validations were done on four published datasets including over 150,000 spectra. The results showed that pMatch can effectively identify unanticipated modifications and significantly increase spectral identification rate.Availabilityhttp://pfind.ict.ac.cn/pmatch/.Supplementary informationSupplementary data are available at Bioinformatics online.
| S-EPMC2881370 | biostudies-literature
Project description:PurposeSerum albumin is in contact with practically all cells in the human body, including tumor cells in cancer patients. The purpose of this study was to explore whether cancer cells affect post-translational modifications (PTMs) of albumin.Material and methodsMass spectrometry was used to identify the PTMs. Purified human serum albumin was incubated with human breast cancer cells MDA-MB-231, MDA-MB-468, MCF7, or kept in water or in cell culture media. PTMs which were affected upon exposure of the albumin to cancer cells were identified. Three-dimensional analysis was performed to locate PTMs in albumin.ResultsWe report here that an exposure to human breast cancer cells affected post-translational modifications (PTMs) of 14 peptides of human serum albumin (HSA). PTMs at 8 peptides were observed upon exposure of HSA to metastatic MDA-MB-231 and MDA-MB-468 breast cancer cells. PTMs at another 6 peptides were lost in MDA-MB-231 and MDA-MB-468 cells, while these 6 PTMs were observed in HSA exposed to conditionally tumorigenic MCF7 cells, or in HSA kept in water or a cell culture medium. Cancer cell altered phosphorylation, deamidation followed by methylation, acetylation, myristylation, palmitoylation, methylation, cysteine persulfide, and S-6-FMN cysteine modifications were detected in HSA. These PTMs locate predominantly in IB and IIA domains of HSA. Three-dimensional analysis showed that this region corresponds to the lipid-binding site and Sudlow's site 1.ConclusionData reported here show that 14 PTMs of human serum albumin can be modified upon its exposure to human breast cancer cells.
| S-EPMC9768621 | biostudies-literature
Project description:BackgroundHistones and their post-translational modifications impact cellular function by acting as key regulators in the maintenance and remodeling of chromatin, thus affecting transcription regulation either positively (activation) or negatively (repression). In this study we describe a comprehensive, bottom-up proteomics approach to profiling post-translational modifications (acetylation, mono-, di- and tri-methylation, phosphorylation, biotinylation, ubiquitination, citrullination and ADP-ribosylation) in human macrophages, which are primary cells of the innate immune system. As our knowledge expands, it becomes more evident that macrophages are a heterogeneous population with potentially subtle differences in their responses to various stimuli driven by highly complex epigenetic regulatory mechanisms.MethodsTo profile post-translational modifications (PTMs) of histones in macrophages we used two platforms of liquid chromatography and mass spectrometry. One platform was based on Sciex5600 TripleTof and the second one was based on VelosPro Orbitrap Elite ETD mass spectrometers.ResultsWe provide side-by-side comparison of profiling using two mass spectrometric platforms, ion trap and qTOF, coupled with the application of collisional induced and electron transfer dissociation. We show for the first time methylation of a His residue in macrophages and demonstrate differences in histone PTMs between those currently reported for macrophage cell lines and what we identified in primary cells. We have found a relatively low level of histone PTMs in differentiated but resting human primary monocyte derived macrophages.ConclusionsThis study is the first comprehensive profiling of histone PTMs in primary human MDM. Our study implies that epigenetic regulatory mechanisms operative in transformed cell lines and primary cells are overlapping to a limited extent. Our mass spectrometric approach provides groundwork for the investigation of how histone PTMs contribute to epigenetic regulation in primary human macrophages.
| S-EPMC4582717 | biostudies-literature