N-terminal Cyclization of Peptides in Large-scale Protein Identification Based on Biological Mass Spectrometry

2009 ◽  
Vol 37 (7) ◽  
pp. 950-954
Author(s):  
Zhuang LU ◽  
Li-Yan ZHAO ◽  
Yang-Jun ZHANG ◽  
Yun CAI ◽  
Yu-Lin DENG ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Large Scale ◽  
Protein Identification ◽  
Biological Mass Spectrometry

Data Mining in Protein Identification by Tandem Mass Spectrometry

2011 ◽  
pp. 472-478
Author(s):  
Haipeng Wang
Keyword(s):  
Mass Spectrometry ◽  
Data Mining ◽  
Tandem Mass Spectrometry ◽  
Large Scale ◽  
Protein Identification ◽  
Peptide Fragmentation ◽  
Tandem Mass ◽  
Fragmentation Pattern ◽  
Post Translational Modification ◽  
Challenges And Opportunities

Protein identification (sequencing) by tandem mass spectrometry is a fundamental technique for proteomics which studies structures and functions of proteins in large scale and acts as a complement to genomics. Analysis and interpretation of vast amounts of spectral data generated in proteomics experiments present unprecedented challenges and opportunities for data mining in areas such as data preprocessing, peptide-spectrum matching, results validation, peptide fragmentation pattern discovery and modeling, and post-translational modification (PTM) analysis. This article introduces the basic concepts and terms of protein identification and briefly reviews the state-of-the-art relevant data mining applications. It also outlines challenges and future potential hot spots in this field.

scholarly journals Ultrafast and Reproducible Proteomics from Small Amounts of Heart Tissue Enabled by Azo and timsTOF Pro

2021 ◽  
Author(s):  
Timothy J Aballo ◽  
David S Roberts ◽  
Jake A Melby ◽  
Kevin M Buck ◽  
Kyle A Brown ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Large Scale ◽  
Protein Identification ◽  
Heart Tissue ◽  
Tryptic Digestion ◽  
Bottom Up ◽  
Preparation Methods ◽  
Liquid Chromatography Tandem Mass ◽  
And Function ◽  
Identification And Quantification

Global bottom-up mass spectrometry (MS)-based proteomics is widely used for protein identification and quantification to achieve a comprehensive understanding of the composition, structure, and function of the proteome. However, traditional sample preparation methods are time-consuming, typically including overnight tryptic digestion, extensive sample clean-up to remove MS-incompatible surfactants, and offline sample fractionation to reduce proteome complexity prior to online liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Thus, there is a need for a fast, robust, and reproducible method for protein identification and quantification from complex proteomes. Herein, we developed an ultrafast bottom-up proteomics method enabled by Azo, a photocleavable, MS-compatible surfactant that effectively solubilizes proteins and promotes rapid tryptic digestion, combined with the Bruker timsTOF Pro, which enables deeper proteome coverage through trapped ion mobility spectrometry (TIMS) and parallel accumulation-serial fragmentation (PASEF) of peptides. We applied this method to analyze the complex human cardiac proteome and identified nearly 4,000 protein groups from as little as 1 mg of human heart tissue in a single one-dimensional LC-TIMS-MS/MS run with high reproducibility. Overall, we anticipate this ultrafast, robust, and reproducible bottom-up method empowered by both Azo and the timsTOF Pro will be generally applicable and greatly accelerate the throughput of large-scale quantitative proteomic studies. Raw data are available via the MassIVE repository with identifier MSV000087476.

scholarly journals Obesity Proteomics: An Update on the Strategies and Tools Employed in the Study of Human Obesity

2018 ◽  
Vol 7 (3) ◽  
pp. 27 ◽  
Author(s):  
Afshan Masood ◽  
Hicham Benabdelkamel ◽  
Assim Alfadda
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
Large Scale ◽  
Protein Identification ◽  
Biochemical Characterization ◽  
Biological Fluids ◽  
Protein Composition ◽  
Cellular Protein ◽  
Human Obesity ◽  
Two Dimensional Gel Electrophoresis

Proteomics has become one of the most important disciplines for characterizing cellular protein composition, building functional linkages between protein molecules, and providing insight into the mechanisms of biological processes in a high-throughput manner. Mass spectrometry-based proteomic advances have made it possible to study human diseases, including obesity, through the identification and biochemical characterization of alterations in proteins that are associated with it and its comorbidities. A sizeable number of proteomic studies have used the combination of large-scale separation techniques, such as high-resolution two-dimensional gel electrophoresis or liquid chromatography in combination with mass spectrometry, for high-throughput protein identification. These studies have applied proteomics to comprehensive biochemical profiling and comparison studies while using different tissues and biological fluids from patients to demonstrate the physiological or pathological adaptations within their proteomes. Further investigations into these proteome-wide alterations will enable us to not only understand the disease pathophysiology, but also to determine signature proteins that can serve as biomarkers for obesity and related diseases. This review examines the different proteomic techniques used to study human obesity and discusses its successful applications along with its technical limitations.

Integrated plant proteomics — putting the green genomes to work

2003 ◽  
Vol 30 (5) ◽  
pp. 471 ◽  
Author(s):  
Joshua L. Heazlewood ◽  
A. Harvey Millar
Keyword(s):  
Mass Spectrometry ◽  
Data Storage ◽  
Large Scale ◽  
Protein Identification ◽  
Mass Spectrometry Analysis ◽  
Plant Science ◽  
Genomic Information ◽  
Plant Proteomics ◽  
Spectrometry Analysis ◽  
Complete Set

Protein analysis has been at the heart of plant science for many years, but with new questions emerging from an abundance of genomic information and further improvements in technology, there are now new opportunities to undertake large-scale analyses and to move to more complex systems than has been possible previously. This explosion of interest and data is often referred to simply as proteomics, which is the study of the complete set of proteins expressed at a given time and place, the proteome. As its name suggests proteomics is intricately linked to allied technologies such as genomics, transcriptomics and metabolomics. In this review of plant proteomics we outline a series of issues that face the practical user, particularly the largest problem that currently faces researchers, the myriad of options to choose from. The choices, problems and pitfalls of entering into gel-based and non-gel-based arraying techniques are discussed together with advances in pre-fractionation of samples, liquid chromatography separations and subcellular analyses. Issues relating to mass spectrometry analysis and the eventual protein identification are outlined, and the dilemmas of data storage and analysis are highlighted. During this tour we provide a series of references to the literature — experimental, theoretical and technical — to illustrate the breadth of current investigations using these techniques.

Microfabricated Modules for Sample Handling, Sample Concentration and Flow Mixing: Application to Protein Analysis by Tandem Mass Spectrometry

1999 ◽  
Vol 121 (1) ◽  
pp. 7-12 ◽  
Author(s):  
D. Figeys ◽  
R. Aebersold
Keyword(s):  
Mass Spectrometry ◽  
Tandem Mass Spectrometry ◽  
Mass Spectrometer ◽  
Dna Sequences ◽  
Large Scale ◽  
Protein Identification ◽  
Protein Analysis ◽  
Tandem Mass ◽  
Modular Systems ◽  
Sample Contamination

The comprehensive analysis of biological systems requires a combination of genomic and proteomic efforts. The large-scale application of current genomic technologies provides complete genomic DNA sequences, sequence tags for expressed genes (EST’s), and quantitative profiles of expressed genes at the mRNA level. In contrast, protein analytical technology lacks the sensitivity and the sample throughput for the systematic analysis of all the proteins expressed by a tissue or cell. The sensitivity of protein analysis technology is primarily limited by the loss of analytes, due to adsorption to surfaces, and sample contamination during handling. Here we summarize our work on the development and use of microfabricated fluidic systems for the manipulation of minute amounts of peptides and delivery to an electrospray ionization tandem mass spectrometer. New data are also presented that further demonstrate the potential of these novel approaches. Specifically, we describe the use of microfabricated devices as modules to deliver femtomole amounts of protein digests to the mass spectrometer for protein identification. We also describe the use of a microfabricated module for the generation of solvent gradients at nl/min flow rates for gradient chromatography-tandem mass spectrometry. The use of microfabricated fluidic systems reduces the risk of sample contamination and sample loss due to adsorption to wetted surfaces. The ability to assemble dedicated modular systems and to operate them automatically makes the use of microfabricated systems attractive for the sensitive and large-scale analysis of proteins.

Identification of BCR-ABL Dependent Gene Regulation by Using a Phosphoproteomics Approach.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2944-2944
Author(s):  
Patrick Ziegler ◽  
Stefan Balabanov ◽  
Ulrike Hartmann ◽  
Winfried Kammer Kammer ◽  
Alfred Nordheim ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Large Scale ◽  
Protein Identification ◽  
Combined Treatment ◽  
K562 Cells ◽  
Peptide Sequencing ◽  
Myelogenous Leukemia ◽  
Imatinib Treatment ◽  
Type Ii ◽  
Cd34 Cells

Abstract Introduction: The selective tyrosine kinase inhibitor imatinib (formerly STI571, Glivecâ) has been shown to block phosphorylation of tyrosine residues by occupying the ATP binding site of the Abl tyrosine kinases Bcr-Abl, c-Abl, v-Abl and Abl-related gene (ARG), as well as platelet-derived growth factor receptor (PDGF) alpha and beta and of the receptor for human stem cell factor (SCF) c-kit. We chose a large scale phospho-proteomics approach to identify novel downstream targets of imatinib which could possibly be utilized for combined treatment strategies. Material and Methods: Phospho-proteomics was performed by comparison of large scale phosphotyrosine-immunoprecipitation of imatinib (10 mM/2 hours) versus DMSO treated K562 cells separated by one-dimensional polyacrylamide gelelectrophoresis. In addition enriched CD34+ cells (>70%) of a newly diagnosed Bcr-Abl positive CML patient were used immediately after purification and treated in the same way as described above. Resulting differentially immuno-precipitated proteins were analyzed using matrix-assisted laser desorption/ionization - time of flight mass spectrometry (MALDI-TOF) and nano electrospray ionization tandem mass spectrometry (ESI-MS/MS). Protein identification via peptide mass-fingerprinting and peptide sequencing was performed using Mascot search tool and NCBI nr database. Differential phosphorylation was confirmed by combined immunoprecipitation and western blot analysis of selected candidate proteins. Results: Phospho-proteomics of K562 cells revealed 8 differentially phosphorylated proteins after a two hour treatment with imatinib including the recently identified c-cbl, and Bcr-Abl itself, the latter confirming autophosphorylation. Ship2 which was originally identified as beeing constitutively phosphorylated in chronic myelogenous leukemia progenitor cells showed reduced, imatinib sensitive phosphorylation. Remaining candidates could be classified as being involved in protein folding or in ATPase activities associated with a variety of functions (type II AAA). The analysis of primary CD34+ cells from a CML patient showed a predominance of the nonmuscular myosin heavy chain protein in different molecular weight forms. Discussion: We detected significant imatinib-dependent differences in protein phosphotyrosine-immunoreactivity of the Bcr-Abl -dependent cell line K562. Previously identified down-stream targets of Bcr-Abl could be confirmed and novel candidate proteins were identified. Phosphorylation of Ship2, a previously identified down-stream target of Bcr-Abl, was found to be inhibited by imatinib treatment. Ongoing studies are aimed at the characterization of the role of the identified phospho-proteins, particularly type II AAAs for Bcr-Abl induced signal transduction as well as for the development of resistance to imatinib.

scholarly journals Stable isotope dimethyl labelling for quantitative proteomics and beyond

2016 ◽  
Vol 374 (2079) ◽  
pp. 20150364 ◽  
Author(s):  
Jue-Liang Hsu ◽  
Shu-Hui Chen
Keyword(s):  
Mass Spectrometry ◽  
Stable Isotope ◽  
Quantitative Proteomics ◽  
Large Scale ◽  
Protein Identification ◽  
Cost Effective ◽  
Quantitative Mass Spectrometry ◽  
Post Translational Modifications ◽  
Labelling Method ◽  
Identification And Characterization

Stable-isotope reductive dimethylation, a cost-effective, simple, robust, reliable and easy-to- multiplex labelling method, is widely applied to quantitative proteomics using liquid chromatography-mass spectrometry. This review focuses on biological applications of stable-isotope dimethyl labelling for a large-scale comparative analysis of protein expression and post-translational modifications based on its unique properties of the labelling chemistry. Some other applications of the labelling method for sample preparation and mass spectrometry-based protein identification and characterization are also summarized. This article is part of the themed issue ‘Quantitative mass spectrometry’.

Algorithm of combining chromatography mass spectrometry-untargeted profiling and multivariate analysis for identification of marker-substances in samples of complex composition

2020 ◽  
Vol 86 (7) ◽  
pp. 12-19
Author(s):  
I. V. Plyushchenko ◽  
D. G. Shakhmatov ◽  
I. A. Rodin
Keyword(s):  
Mass Spectrometry ◽  
Multivariate Analysis ◽  
Large Scale ◽  
Complex Composition ◽  
Unified Protocol ◽  
Chromatography Mass Spectrometry ◽  
Marker Substances ◽  
Selection Testing ◽  
Untargeted Profiling

A viral development of statistical data processing, computing capabilities, chromatography-mass spectrometry, and omics technologies (technologies based on the achievements of genomics, transcriptomics, proteomics, metabolomics) in recent decades has not led to formation of a unified protocol for untargeted profiling. Systematic errors reduce the reproducibility and reliability of the obtained results, and at the same time hinder consolidation and analysis of data gained in large-scale multi-day experiments. We propose an algorithm for conducting omics profiling to identify potential markers in the samples of complex composition and present the case study of urine samples obtained from different clinical groups of patients. Profiling was carried out by the method of liquid chromatography mass spectrometry. The markers were selected using methods of multivariate analysis including machine learning and feature selection. Testing of the approach was performed using an independent dataset by clustering and projection on principal components.

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