Author: Jeremy Lynn BalsbaughPublisher:
ISBN:
Category :
Languages : en
Pages :
Book Description
Studies in Protein Post-translational Modification Using CAD and ETD Mass Spectrometry
Analysis of Protein Post-Translational Modifications by Mass Spectrometry
Author: John R. GriffithsPublisher: John Wiley & Sons
ISBN: 1119045851
Category : Science
Languages : en
Pages : 414
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
Studies of Protein Post-translational Modifications Using High Resolution Tandem Mass Spectrometry
Author: Huilin LiETD and ECD Mass Spectrometry Fragmentation for the Characterization of Protein Post Translational Modifications
Author: Lisa ElviriPublisher:
ISBN: 9789535101413
Category :
Languages : en
Pages :
Book Description
Post-translational Modifications of Proteins
Author: Christoph KannichtPublisher: Humana Press
ISBN: 9781588297198
Category : Science
Languages : en
Pages : 0
Book Description
This is a fully up-dated and expanded practical guide to protein structure-function relationships. This important area of research is brought up-to-date by the leading scientists in the field. The compilation of detailed protocols focuses on protein function, proteome research and characterization of pharmaceutical proteins, while following the successful format of the Methods in Molecular BiologyTM series. Comprehensive and cutting edge, the book serves as practical guide for researchers working in the field of protein structure-function relationships and the rapidly growing field of proteomics, as well as scientists in the pharmaceutical industries.
Post-Translational Modification of Proteins
Author: Christoph KannichtPublisher: Humana
ISBN: 9781493990535
Category : Science
Languages : en
Pages : 0
Book Description
The majority of all proteins undergo co- and/or post-translational modifications, crucially altering physical and chemical properties, folding, conformation distribution, stability, activity, and, consequently, the function of the proteins. In Post-Translational Modifications of Proteins: Tools for Functional Proteomics, Third Edition this important area of research is brought up-to-date by the leading scientists in the field. This compilation of detailed protocols focuses on protein function, proteome research and characterization of pharmaceutical proteins, while following the successful format of the Methods in Molecular Biology series. Each chapter provides a brief introduction to the topic, step-by-step laboratory protocols, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls. Comprehensive and cutting edge, Post-Translational Modifications of Proteins: Tools for Functional Proteomics, Third Edition serves as practical guide for researchers working in the field of protein structure-function relationships and the rapidly growing field of proteomics, as well as scientists in the pharmaceutical industries.
Posttranslational Modification of Proteins
Author: Christoph KannichtPublisher: Humana Press
ISBN: 9780896036789
Category : Science
Languages : en
Pages : 322
Book Description
Christoph Kannicht and a panel of highly experienced researchers describe readily reproducible methods for detecting and analyzing the posttranslational modifications of protein, particularly with regard to protein function, proteome research, and the characterization of pharmaceutical proteins. Among the methods presented are those for analyzing the assignment of disulfide bond sites in proteins, protein N-glycosylation and protein O-glycosylation, and oligosaccharides present at specific single glycosylation sites in a protein. Additional powerful techniques facilitate the analysis of glycosylphosphatidylinositols, lipid modifications, protein phosphorylation and sulfation, protein methylation and acetylation, a-amidation, g-glutamate, isoaspartate, and lysine hydroxylation.
Advancing Quantitative Proteomics and Protein Post-translational Modification Analyses by Multi-dimensional Mass Spectrometric Approaches
Author: Zihui LiPublisher:
ISBN:
Category :
Languages : en
Pages : 0
Book Description
Proteins are important molecules that are involved in numerous critical biological processes. Consequently, understanding the functional roles and dysregulation of proteins constitutes a vital task in both basic and clinical research. However, it is very challenging to define the full complement of proteins due to their large degree of variations, especially considering that many proteins can undergo numerous post-translational modifications (PTMs) which dramatically affect protein conformations and functions. Recently, mass spectrometry (MS) has evolved as a powerful tool to characterize proteins and their PTMs benefiting from its high speed, sensitivity and the ability to monitor thousands of analytes simultaneously. This dissertation is devoted to the development and application of various MS-based strategies to facilitate the detection of proteins and their PTMs qualitatively and quantitatively. A portion of this dissertation describes the investigation of proteome-wide alterations with the development and maturation of human pancreas using custom-developed isobaric tags. This study focuses on the extracellular matrix proteins and provides information on the localization changes of selected proteins in islet or acinar compartments as well. The normal baseline data across the lifespan of human pancreas will become a valuable resource and benefit future studies into the pancreatic pathologies. Switching gear to protein PTMs, this dissertation documents the design and development of a novel biotin thiol tag to greatly improve the in-depth profiling of two important PTMs, citrullination and homocitrullination, from complex biological samples. The utility of the approach is demonstrated by providing the first tissue-specific citrullination and homocitrullination atlas in mice. The first high-throughput quantitative analysis pipeline of protein citrullination is subsequently developed by the integration with isobaric labeling strategy. We also further improve the protein citrullination detection by using a combination of various digestion and fragmentation techniques. Citrullination alteration during the progression of Alzheimer's disease is further explored using human cerebrospinal fluid to shed light on the disease pathogenesis. This dissertation also highlights the application of a novel sub-atmospheric pressure matrix-assisted laser desorption/ionization mass spectrometry platform for the in situ imaging of N-glycans. Spatial distribution of N-glycans in ovarian cancer tissue section may indicate potential association of this modification with tumor progression. Collectively, we expect the biological insights and technological advancements presented in this work will lead to improved understanding of microenvironment in both healthy and disease conditions, and the new knowledge gained from these studies will inspire and benefit future research effort in related fields.
Characterization and Identification of Protein Posttranslational Modifications Using Protein Enrichment and Mass Spectrometry
Author: Liwen WangPublisher:
ISBN:
Category :
Languages : en
Pages :
Book Description
Abstract: This dissertation describes a proteomic workflow for the analysis of protein post-translational modifications (PTMs). The workflow combines the techniques for protein enrichment, multi-dimensional separations, mass spectrometry (MS) and automatic data analysis. The workflow was developed to improve the application of proteomic analysis in the realms of biomarker discovery and experimental therapeutic research. Chapter 2 presents an immunoaffinity chromatography method that was developed to enrich acetylated histones. A self-packed immunoaffinity capillary column was developed using commercial antibodies that could be recycled and used for on-line and off-line enrichment. The acetylated fractions were collected and identified by Matrix Assisted Laser Desorption (MALDI) MS and electrospray ionization (ESI) liquid chromatography tandem mass spectrometry (LC-MS/MS). In chapter 3 an optimized phosphoproteomic analysis workflow based on phosphopeptide enrichment, data-dependant neutral loss mass spectrometry and a novel hierarchical database searching is described. The combination of these approaches improved the confidence of phosphopeptide identifications. Chapter 4 describes the use of phosphoprotein enrichment and a tandem phosphoprotein and phosphopeptide enrichment to improve the identification of phosphoproteins and localization of the phosphorylation sites. Purification of global phosphoproteins from primary CLL B-cells was conducted by use of PhosTag Zn2 enrichment strategy at neutral pH. SDS-PAGE gel was used to separate the purified phosphoprotein fraction and Pro-Q diamond staining was employed to visualize those phosphoprotein bands. Shot-gun proteomic analysis was then performed to identify all the enriched phosphoproteins in the gel. Phosphopeptide enrichment was used in tandem to map phosphorylation sites of the enriched phosphoproteins. Chapter 5 describes the identification of tyrosine phosphoproteins associated with immunotherapy of malignant cells with the small modular immunopharmaceutical targeted against CD37 (CD37-SMIPTM). This drug induces apoptosis and antibody-dependent cellular cytotoxicity (ADCC) in primary Chronic Lymphocyte Leukemia (CLL) cells. Tyrosine phosphorylation of proteins was investigated as an early activation event for the cytotoxicity. Immunoprecipitation was used to purify the phosphotyrosine proteins from treated cell lysate and untreated cell lysate. Detection of modulation of tyrosine phosphorylation and identification of those tyrosine phosphoproteins after treatment by proteomic approaches revealed proteins associated with the signaling pathway activated by immunotherapy. Chapter 6 describes a direct application of the proteomic platform developed in Chapter 3 combined with LC-MS protein profiling. The modulation of histone phosphorylation isoforms induced by various chemotherapy drugs was detected by LC-MS screening. We detected the dephosphorylation of histones H1 and hyperphosphorylation of H2A.X associated with the different drug treatments.
Mass Spectrometric Investigations of Hidden Post-translational Modifications and the Inevitable Fate of Long-lived Proteins
Author: Yana Alexandra LyonPublisher:
ISBN: 9780438897458
Category : Mass spectrometry
Languages : en
Pages : 154
Book Description
Protein turnover is essential for the vitality of a functional proteome, as it allows for old and damaged proteins to be destroyed and replaced with newly synthesized copies. However, there is a growing class of proteins that escape this process and are classified as long-lived proteins (LLPs). The intrinsic instability of LLPs makes them susceptible to damage through the accumulation of unwanted post-translational modifications (PTMs), and mounting evidence suggests that this plays a critical role in aging and disease. It is known that crystallin turnover essentially stops in the mature lens fiber cells of the eye which makes it an ideal target for understanding damage due to aging, however mapping out the ensuing degradation has proven to be a formidable challenge. There are a myriad of spontaneous PTMs that accrue in the lens, including isomerization and epimerization, both of which are invisible to many traditional analytical techniques. Such small perturbations to individual amino acids may appear to be innocuous, but in fact, these hidden modifications impact protein structure, function, and solubility, all of which are crucial for lens transparency. The details of protein aging are examined herein by employing a novel mass spectrometric dissociation technique that utilizes radical chemistry to detect sites of isomerization and epimerization. Studies on young animal eye lenses reveal how the degree of these modifications change as a function of protein solubility and shows that specific structural motifs and amino acid repeats serve as isomerization hotspots. Investigations on human donor lenses expose an increased presence of isoAspartyl residues in the nucleus of the lens compared to the cortex, suggesting decreased activity of a specific repair enzyme. Delving deeper into the structural and functional consequences of these modifications required the use of native mass spectrometry, enzyme assays, and molecular dynamics calculations, all of which help show that proper crystallin oligomerization can be disrupted by these PTMs. An alternative approach to RDD utilizing electron transfer dissociation (ETD) was also developed to detect epimerized residues in peptide models. Collectively, this work serves to better our understanding of how isomerization and epimerization contribute to the inevitable fate of LLPs.