Advancing Electron Transfer Dissociation Technologies for Characterization of Proteomes and Post-translational Modifications PDF Download

Author: Nicholas M. Riley
Publisher:
ISBN:
Category :
Languages : en
Pages : 0

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Book Description
This dissertation presents research focusing on the development of new instrumentation and methodology to leverage ion-ion reactions for proteomic analyses. Electron transfer dissociation (ETD) technologies have proven a valuable alternative to collision-based fragmentation methods for sequencing peptides and proteins to advance global proteome characterization. Chapter 1 outlines the core concepts central to mass spectrometry (MS)-based proteomics, in addition to the basic principles of ETD and various strategies to improve its efficacy - including the technology that is the focus of this work, i.e., activated ion ETD (AI-ETD). Chapter 2 describes the first application of AI-ETD to intact proteins, which are more chemically complex and, thus, more difficult to sequence, than their peptide counterparts. Chapter 3 discusses a new strategy to improve signalto- noise in ETD spectra, which is especially beneficial for intact protein analysis and which has been incorporated into the newest generation of commercially available quadrupole-Orbitrap-linear ion trap hybrid MS systems. AI-ETD capabilities were also recently implemented on this stateof- the-art MS system (Chapter 4), and the ability to perform AI-ETD on this instrument enables comprehensive sequence coverage of moderately-sized intact proteins (Chapter 5), significantly improves proteoform characterization in large-scale analyses of complex mixtures of intact proteins (Chapter 6), and also enhances characterization of larger intact proteins (Chapter 7). Furthermore, AI-ETD improves characterization of post-translational modifications. Chapter 8 demonstrates the utility of AI-ETD for phosphosite localization in phosphopeptides and intact phosphoproteins, and Chapter 9 presents the largest glycoproteomic study to date by using AI-ETD to interrogate intact N-glycopeptides. Beyond positive-mode analyses of peptide and protein cations, ion-ion reactions also bring unique benefits to negative-mode analyses of precursor anions, where collision-based dissociation fails to consistently produce sequence-informative fragments. Chapter 10 describes implementation of negative ETD (NETD) and activated ion NETD (AI-NETD) and their application to whole-proteome sequencing in the negative mode, and Chapter 11 presents a modified search algorithm to improve interpretation of large-scale NETD and AI-NETD data. Conclusions and future directions of these projects are discussed in Chapter 12.

Author: Trenton M. Peters-Clarke
Publisher:
ISBN:
Category :
Languages : en
Pages : 0

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This dissertation details innovations in mass spectrometry-based instrumentation that expand our ability to survey biopolymers, including proteins, peptides, and oligonucleotides, with increased sensitivity, speed, and depth. Chapter 1 provides an overview of mass spectrometry instrumentation at the leading edge of proteomics and for nucleic acid analysis. In Chapter 2, the optical-fiber enabled coupling of a mass spectrometer to an infrared laser improves proteomic characterization. For Chapter 3, electron transfer dissociation-based proteomics is investigated and inefficiencies of related ion-ion reactions are characterized. Chapter 4 introduces a novel nucleic acid sequencing technology with activated ion-negative electron transfer dissociation to profile RNA molecules and their modifications. In Chapter 5, tandem mass spectrometry comprehensively characterizes heavily-modified RNA therapeutics. Chapter 6 focuses on a proteome-wide analysis of the bacterial post-translational modification O-mycoloylation. In Chapter 7, a novel tandem mass spectrometry strategy, termed activated ion-tandem mass tags, is revealed, boosting quantitative reporter ion generation. Chapter 8 expands upon AI-TMT for quantitative single-cell proteomics, realizing sensitivity gains which drive a greater dynamic range of quantification. In Chapter 9, a novel mass spectrometer enables the one hour human proteome and we explore one minute proteome analyses, pushing the limits of high-throughput proteome analysis. Chapter 10 discusses emergent proteomic techniques and suggests future directions of the field.

Author: Ying Ge
Publisher:
ISBN:
Category :
Languages : en
Pages : 354

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Author: John R. Griffiths
Publisher: John Wiley & Sons
ISBN: 1119045851
Category : Science
Languages : en
Pages : 414

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Book Description
Covers all major modifications, including phosphorylation, glycosylation, acetylation, ubiquitination, sulfonation and and glycation Discussion of the chemistry behind each modification, along with key methods and references Contributions from some of the leading researchers in the field A valuable reference source for all laboratories undertaking proteomics, mass spectrometry and post-translational modification research

Author: Jean-Pierre Schermann
Publisher: Elsevier
ISBN: 0080558224
Category : Science
Languages : en
Pages : 499

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Book Description
Spectroscopy and Modeling of Biomolecular Building Blocks presents an overview of recent advances in the intertwining of the following research fields: photon and electron spectroscopy, quantum chemistry, modelling and mass-spectrometry. The coupling of these disciplines offers a new point of view to the understanding of isolated elementary building blocks of biomolecules and their assemblies. It allows the unambiguous separation between intrinsic properties of biomolecular systems and those induced by the presence of their environment. The first chapters provide background in modelling (I), frequency-resolved spectroscopy using microwave, infrared and UV photons, time-resolved spectroscopy in the femtosecond domain and energy-resolved electron spectroscopy (II) and production of gas-phase neutral and ionic biomolecular species, mass-spectrometry, ion mobility and BIRD techniques (III). Chapter IV is devoted to case studies of gas-phase experimental investigations coupled to quantum or classical calculations. The topics are structural studies of nucleobases and oligonucleotides, peptides and proteins, sugars; neuromolecules; non-covalent complexes; chiral systems, interactions of low-energy electrons with biomolecules in the radiation chemistry context and very large gas-phase biomolecular systems. The fifth chapter concerns the link between gas-phase and liquid-phase. Different treatments of solvation are illustrated through examples pointing out the influence of progressive addition of water molecules upon properties of nucleobases, peptides, sugars and neuromolecules. Offer a new perspective to the understanding of isolated elementary building blocks of bio molecules Includes case studies of experimental investigations coupled to quantum or classical calculations

Author: Conrad Bessant
Publisher: Royal Society of Chemistry
ISBN: 1782626735
Category : Science
Languages : en
Pages : 429

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Book Description
The field of proteomics has developed rapidly over the past decade nurturing the need for a detailed introduction to the various informatics topics that underpin the main liquid chromatography tandem mass spectrometry (LC-MS/MS) protocols used for protein identification and quantitation. Proteins are a key component of any biological system, and monitoring proteins using LC-MS/MS proteomics is becoming commonplace in a wide range of biological research areas. However, many researchers treat proteomics software tools as a black box, drawing conclusions from the output of such tools without considering the nuances and limitations of the algorithms on which such software is based. This book seeks to address this situation by bringing together world experts to provide clear explanations of the key algorithms, workflows and analysis frameworks, so that users of proteomics data can be confident that they are using appropriate tools in suitable ways.

Author: Sanjeeva Srivastava
Publisher: Springer
ISBN: 8132228375
Category : Medical
Languages : en
Pages : 120

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Book Description
This book is oriented towards post-graduates and researchers with interest in proteomics and its applications in clinical biomarker discovery pipeline. Biomarker discovery has long been the research focus of many life scientists globally. However, the pipeline starting from discovery to validation to regulation as a diagnostic or therapeutic molecule follows a complex trajectory. This book aims to provide an in-depth synopsis on each of these developmental phases attendant to biomarker “life cycle” with emphasis on the emerging and significant role of proteomics. The book begins with a perspective on the role of biorepositories and need for biobanking practices in the developing world. The next chapter focuses on disease heterogeneity in context to geographical bias towards susceptibility to the disease and the role of multi-omics techniques to devise disruptive innovations towards biomarker discovery. Chapter 3 focuses on various omics-based platforms that are currently being used for biomarker discovery, their principles and workflow. Mass spectrometry is emerging as a powerful technology for discovery based studies and targeted validation. Chapter 4 aims at providing a glimpse of the basic workflow and considerations in mass spectrometry based studies. Rapid and aptly targeted research funding has often been deemed as one of the decisive factors enabling excellent science and path breaking innovations. With the need for sophistication required in multi-omics research, Chapter 5 focuses on innovative funding strategies such as crowdfunding and Angel philanthropy. Chapter 6 provides the latest advances in education innovation, the premise and reality of bioeconomy especially in a specific context of the developing world, not to mention the new concept of “social innovation” to link biomarkers with socially responsible and sustainable applications. Chapter 7, in ways similar to biomarkers, discusses the biosimilars as a field that has received much focus and prominence recently due to their immense potential in clinical and pharmaceutical innovation literatures. The broader goal post-biomarker discovery is to translate their use in clinics. However, the road from bench-to-bed side is arduous and complex that is subject to oversight from various national and international regulatory bodies. Chapter 8 underscores these regulatory science considerations and provides a concise overview on intellectual property rights in biomarker discovery. Thus, this book contributed by eminent biomarker scientists, clinicians, translational researchers and social scientists holistically covers the various facets of the biomarker discovery journey from “cell to society” in developing world. The lessons learned and highlighted here are of interest to the life sciences community in a global and interdependent world.

Author: Nadezda Sargaeva
Publisher:
ISBN:
Category :
Languages : en
Pages : 354

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Book Description
Abstract: Electron capture dissociation (ECD) and electron transfer dissociation (ETD) can generate unique fragments and preserve post-translational modifications (PTMs), enabling their detection in biological samples. They have been used to differentiate isomeric aspartic (Asp) and isoaspartic acids (isoAsp) produced upon non-enzymatic deamidation of asparagine (Asn) -- a frequently occurring PTM. IsoAsp formation was detected in amyloid-B (AB) peptides in the specimens of Alzheimer's disease (AD) patients, and is a potential biomarker for AD if it can be detected early in biofluids of live individuals. Synthetic isoAsp-containing AB fragments were studied using ECD to test the method's applicability. IsoAsp-7 and -23 were detected in top-down analysis of the 4.5kDa AB42 protein and in AB17-28 peptide. Further, a related method, electron ionization/impact dissociation (EID), was successfully applied to Asp/isoAsp differentiation for the first time. High-performance liquid chromatography (HPLC) is a powerful technique for the separation of complex mixtures. HPLC separation of Asp- and isoAsp-containing peptides revealed inconsistent elution orders, especially when isoAsp was located at the N-terminus, requiring ECD for identification. New diagnostic fragments were proposed for N-terminal isoAsp based on the ECD and ETD results. Challenges in detection of such fragments were improved by supplemental activation and chemical modifications. Furthermore, a model for retention time prediction was applied to isoAsp-containing peptides and suggested for their improved identification in HPLC-MS/MS approach. IsoAsp is a B-amino acid, which distinctively contains an additional methylene group in the backbone, forming a C a -CB bond. Cleavage of this bond provides diagnostic fragments for isoAsp by ECD. The same was proposed for other B-amino acids. However, the Ca -CB ; bond cleavages were rare due to the instability of the CB radical. Alternatively, in-source decay (ISD) fragmentation during matrix-assisted laser desorption/ionization (MALDI) process can produce abundant ECD-like fragmentation. It was proposed that use of hydrogen-accepting matrices may lead to Ca -CB bond cleavage in B-amino acids, because the resulting radical would be stabilized by the carbonyl group. To test this, B-amino acid-containing peptides were analyzed by MALDI-ISD using 5-nitrosalicylic acid matrix. The Ca -CB bond cleavages were observed. Overall, new and improved methods were implemented allowing better characterization and differentiation of B-amino acids.

Author: Xianquan Zhan
Publisher: BoD – Books on Demand
ISBN: 1838800336
Category : Science
Languages : en
Pages : 92

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Book Description
A proteoform is the basic unit in a proteome, defined as its amino acid sequence + post-translational modifications + spatial conformation + localization + cofactors + binding partners + a function, which is the final functional performer of a gene. Studies on proteoforms offer in-depth insights and can lead to the discovery of reliable biomarkers and therapeutic targets for effective prediction, diagnosis, prognostic assessment, and therapy of disease. This book focuses on the concept, study, and applications of proteoforms. Chapters cover such topics as methodologies for identifying and preparing proteoforms, proteoform pattern alteration in pituitary adenomas, and proteoforms in leukemia.

Author:
Publisher: Academic Press
ISBN: 0128186704
Category : Science
Languages : en
Pages : 648

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Book Description
Post-translational Modifications That Modulate Enzyme Activity, Volume 626 in the Methods in Enzymology series, continues the legacy of this premier serial with quality chapters authored by leaders in the field. Updated chapters include Crosstalk between cellular metabolism and histone acetylation, Isolation of protein complexes and modifications that regulate transcriptional machinery, High-throughput phosphoproteome mapping through multiplexed mass spectrometry, Differentiation of D and L epimerization in proteins, Biochemical analysis of protein arginylation, Site-specific Determination of lysine acetylation stoichiometries on the proteome-scale, Genomic and biochemical analysis of RNA post-transcriptional modifications, Isolation and characterization of glycosylated (neuro)peptides, and more. Provides the authority and expertise of leading contributors from an international board of authors Presents the latest release in the Methods in Enzymology series Includes the latest information on Post-translational Modifications that Modulate Enzyme Activity