Abstract:
:Affinity purification coupled to mass spectrometry (AP-MS) is gaining widespread use for the identification of protein-protein interactions. It is unclear, however, whether typical AP sample complexity is limiting for the identification of all protein components using standard one-dimensional LC-MS/MS. Multidimensional sample separation is useful for reducing sample complexity prior to MS analysis and increases peptide and protein coverage of complex samples. Here, we monitored the effects of upstream protein or peptide separation techniques on typical mammalian AP-MS samples, generated by FLAG affinity purification of four baits with different biological functions and/or subcellular distribution. As a first separation step, we employed SDS-PAGE, strong cation exchange LC, or reversed-phase LC at basic pH. We also analyzed the benefits of using an instrument with a faster scan rate, the new TripleTOF 5600 mass spectrometer. While all multidimensional approaches yielded a clear increase in spectral counts, the increase in unique peptides and additional protein identification was modest and came at the cost of increased instrument and handling time. The use of a high duty-cycle instrument achieved similar benefits without these drawbacks. An increase in spectral counts is beneficial when data analysis methods relying on spectral counts, including Significance Analysis of INTeractome (SAINT), are used.
journal_name
Proteomicsjournal_title
Proteomicsauthors
Dunham WH,Larsen B,Tate S,Badillo BG,Goudreault M,Tehami Y,Kislinger T,Gingras ACdoi
10.1002/pmic.201000571subject
Has Abstractpub_date
2011-07-01 00:00:00pages
2603-12issue
13eissn
1615-9853issn
1615-9861journal_volume
11pub_type
杂志文章相关文献
PROTEOMICS文献大全abstract::To better understand the underlying molecular basis of leaf development in maize, a reference map of nuclear proteins in basal region of seedling leaf was established using a combination of 2DE and MALDI-TOF-MS. In total, 441 reproducible protein spots in nuclear proteome of maize leaf basal region were detected with ...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201300147
更新日期:2014-05-01 00:00:00
abstract::In theory, proteases with broad cleavage specificity could be applied to digest protein samples to improve the phosphoproteomic analysis coverage. However, in practice this approach is seldom employed. This is because the identification of phosphopeptides without enzyme specificity by conventional database search stra...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201400268
更新日期:2014-11-01 00:00:00
abstract::Sporozoites of the malaria parasite Plasmodium are transmitted by mosquitoes and infect the liver for an initial and obligatory round of replication, before exponential multiplication in the blood and onset of the disease. Sporozoites and liver stages provide attractive targets for malaria vaccines and prophylactic dr...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.202000305
更新日期:2021-01-16 00:00:00
abstract::Puccinia triticina causes leaf rust, a disease that causes annual yield losses in wheat. It is an obligate parasite that invades the host leaf and forms intracellular structures called haustoria, which obtain nutrients and suppress host immunity using secreted proteins called effectors. Since effector proteins act at ...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201400241
更新日期:2015-04-01 00:00:00
abstract::Identification and characterization of biomarkers in body fluids such as serum or urine serve as a basis for early detection of diseases, particularly of cancer. Performing 2-DE with subsequent MS analyses, conventional immunoblotting and immunohistochemistry we identified two proteins, orosomucoid (ORM) and human zin...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200402005
更新日期:2005-11-01 00:00:00
abstract::High temperatures during grain filling have been reported to be one of the factors that can affect the dough properties and quality characteristics of wheat. Responses to high temperature have been related to changes in protein composition at both quantitative and qualitative levels. The present study was conducted to...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200390026
更新日期:2003-02-01 00:00:00
abstract::We describe the first phosphoproteome of the model filamentous fungus Aspergillus nidulans. Phosphopeptides were enriched using titanium dioxide, separated using a convenient ultra-long reverse phase gradient, and identified using a "high-high" strategy (high mass accuracy on the parent and fragment ions) with higher-...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201400063
更新日期:2014-11-01 00:00:00
abstract::Heat shock proteins (HSPs) are crucial for the maintenance of cell integrity during normal cellular growth as well as during pathophysiological conditions. While functioning mainly as molecular chaperones, HSPs also appear to be involved in diverse biological activities, such as apoptosis, carcinogenesis, and cytoprot...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200400951
更新日期:2004-11-01 00:00:00
abstract::Caulobacter crescentus, a Gram negative alpha-purple bacterium that displays an invariant asymmetric cell division pattern, has become a key model system for the study of bacterial development. Membrane proteins play key roles in cell cycle events, both as components of landmark morphological structures and as critica...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/1615-9861(200104)1:5<705::AID-PROT705>3.0.
更新日期:2001-05-01 00:00:00
abstract::Previous studies have found that some first onset schizophrenia patients show signs of impaired insulin signaling. Also, epidemiological studies have shown that periods of suboptimal nutrition including protein deficiencies during pregnancy can lead to increased incidence of metabolic conditions and psychiatric disord...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201200376
更新日期:2012-12-01 00:00:00
abstract::Attribution of the most probable functions to proteins identified by proteomics is a significant challenge that requires extensive literature analysis. We have developed a system for automated prediction of implicit and explicit biologically meaningful functions for a proteomics study of the nucleolus. This approach u...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200600693
更新日期:2007-03-01 00:00:00
abstract::A proteomic analysis of a wild-type and of a phoB mutant showed that Vibrio cholerae expresses genes of two major regulons in response to phosphate starvation. The Pho regulon, expressed by the wild-type, allowed the cells to adapt to the new environment. Induction of the general stress regulon was mainly observed in ...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200500238
更新日期:2006-03-01 00:00:00
abstract::Tectona grandis L. (teak) is one of the premier hardwood timbers in the world, ranking at present in the top five tropical hardwood species in terms of worldwide plantation area. Characterization of the proteins present in teak leaves will provide a basis for the development of new tools aimed at assisting tree select...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201100183
更新日期:2012-04-01 00:00:00
abstract::Changing smooth muscle phenotype and abnormal cell proliferation are important features of vascular pathology, including the failure of saphenous vein bypass grafts. We have characterised and mapped protein expression in human saphenous vein medial smooth muscle, using two-dimensional (2-D) polyacrylamide gel electrop...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/1615-9861(200111)1:11<1405::AID-PROT1405>3
更新日期:2001-11-01 00:00:00
abstract::The protein components of urine are useful indicators of renal function and human health in general. Urine samples are easily attainable making them ideal substrates for biomarker research. Analysis of the urine proteome however, has been hindered by the great variability of the urine specimens, and the presence of va...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200500671
更新日期:2006-08-01 00:00:00
abstract::Glycan-binding proteins (GBPs) play an important role in cell adhesion, bacterial/viral infection, and cellular signaling pathways. However, little is known about the precision alteration of GBPs referred to pathological changes in hepatic stellate cells (HSCs) during liver fibrosis. Here, the carbohydrate microarrays...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201500030
更新日期:2015-10-01 00:00:00
abstract::Protein arrays permit the parallel analysis of many different markers in a small sample volume. However, the problem of cross-reactivity limits the degree of multiplexing in parallel sandwich immunoassays (using monoclonal antibodies (mAbs)), meaning antibodies must be prescreened in order to reduce false positives. I...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200300736
更新日期:2004-05-01 00:00:00
abstract::We describe a new approach for the characterization of a digested protein complex with quantitative aspects. Accurate masses of tryptic peptides in the digested complex were acquired by nano-liquid chromatography Fourier transform-ion cyclotron resonance mass spectrometry (MS). The conditions of the electrospray ion s...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200300812
更新日期:2004-09-01 00:00:00
abstract::Glycosylation is a widespread PTM of proteins; the carbohydrate moieties provide various functional, immunological and structural aspects of both eukaryotic and prokaryotic glycoproteins. Traditional strategies used to analyse glycoprotein O-glycans involve glycoprotein isolation, followed by glycan release using solu...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200500331
更新日期:2006-05-01 00:00:00
abstract::We report on a novel strategy for identification of specific sulfation motifs in chondroitin/dermatan sulfate (CS/DS) chain derived from decorin (Dcn), based on enzyme cleavage and multistage MS (MS(n)). Released CS/DS chains were digested with chondroitin B and in parallel with AC I lyases to obtain oligosaccharides ...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200800440
更新日期:2009-07-01 00:00:00
abstract::Two-dimensional difference gel electrophoresis (2-D DIGE) coupled with mass spectrometry (MS) was used to investigate tumor-specific changes in the proteome of human colorectal cancers and adjacent normal mucosa. For each of six patients with different stages of colon cancer, Cy5-labeled proteins isolated from tumor t...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200300635
更新日期:2004-03-01 00:00:00
abstract::Gene expression analyses of embryonic stem cells (ESCs) will help to uncover or further define signaling pathways and molecular mechanisms involved in the maintenance of self-renewal and pluripotency. We employed a 2-DE-based proteomics approach to analyze human ESC line, Royan H5, in undifferentiated cells and differ...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200900003
更新日期:2009-11-01 00:00:00
abstract::Previously, we reported that nucleophosmin (NPM) was increased in glioblastoma multiforme (GBM). NPM is a phosphoprotein related to apoptosis, ribosome biogenesis, mitosis, and DNA repair, but details about its function remain unclear. We treated U87MG and A172 cells with small interference RNA (siRNA) and obtained a ...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201200034
更新日期:2012-08-01 00:00:00
abstract::Previous investigations of bovine seminal plasma (BSP) have revealed the identities of the three major proteins, BSP-PDC109, BSP-A3 and BSP-30 kDa, which together constitute about half of the total protein, as well as about 30 of the minor proteins. Analyses of BSP by 2-DE have revealed about 250 protein spots, sugges...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200500830
更新日期:2006-11-01 00:00:00
abstract::Protein quantification at proteome-wide scale is an important aim, enabling insights into fundamental cellular biology and serving to constrain experiments and theoretical models. While proteome-wide quantification is not yet fully routine, many datasets approaching proteome-wide coverage are becoming available throug...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201400441
更新日期:2015-09-01 00:00:00
abstract::A brief period of ischemia followed by timely reperfusion may lead to prolonged, yet reversible, contractile dysfunction (myocardial stunning). Damage to the myocardium occurs not only during ischemia, but also during reperfusion, where a massive release of oxygen-free radicals (OFR) occurs. We have previously utilize...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200600219
更新日期:2006-12-01 00:00:00
abstract::A proteomics approach has been used to study changes in protein abundance during leaf senescence in white clover. Changes in cell ultrastructure were also examined using transmission electron microscopy. The most obvious ultrastructural changes during senescence occurred in chloroplasts, with progressive loss of thyla...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/1615-9861(200209)2:9<1114::AID-PROT1114>3.
更新日期:2002-09-01 00:00:00
abstract::In the era of quantitative proteomics, where mass spectrometry plays a pivotal role, in particular associated with the use of data-independent acquisition, it is time to perform an overview of this growing field with special focus on one of the most promising approaches: SWATH-MS, and to present future perspectives fo...
journal_title:Proteomics
pub_type: 杂志文章,评审
doi:10.1002/pmic.201600278
更新日期:2017-02-01 00:00:00
abstract::Collision-activated dissociation and electron-transfer dissociation (ETD) each produce spectra containing unique features. Though several database search algorithms (e.g. SEQUEST, MASCOT, and Open Mass Spectrometry Search Algorithm) have been modified to search ETD data, this consists chiefly of the ability to search ...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.200900570
更新日期:2010-01-01 00:00:00
abstract::The PIK3CA gene encodes for the p110 alpha isoform of PI3 kinase and is one of the most frequently mutated oncogenes in human cancers. However, the mechanisms by which PIK3CA mutations activate cell signaling are not fully understood. Here we used a phosphoproteomic approach to compare differential phosphorylation pat...
journal_title:Proteomics
pub_type: 杂志文章
doi:10.1002/pmic.201400342
更新日期:2015-01-01 00:00:00