A Study of Protein Interactions by Electrospray Ionization Mass Spectrometry PDF Download
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Author:
Publisher:
ISBN:
Category : Mass spectrometry
Languages : en
Pages :
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Author:
Publisher:
ISBN:
Category : Mass spectrometry
Languages : en
Pages :
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Author: Kevin Downard
Publisher: John Wiley & Sons
ISBN: 047014632X
Category : Science
Languages : en
Pages : 153
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Book Description
The authoritative guide to analyzing protein interactions by mass spectrometry Mass spectrometry (MS) is playing an increasingly important role in the study of protein interactions. Mass Spectrometry of Protein Interactionspresents timely and definitive discussions of the diverse range of approaches for studying protein interactions by mass spectrometry with an extensive set of references to the primary literature. Each chapter is written by authors or teams of authors who are international authorities in their fields. This leading reference text: * Discusses the direct detection of protein interactions through electrospray ionization (ESI-MS); ion mobility analysis; and matrix-assisted laser desorption/ionization (MALDI-MS) * Covers the indirect analysis of protein interactions through hydrogen-deuterium exchange (HX-MS); limited proteolysis; cross-linking; and radial probe (RP-MS) * Guides researchers in the use of mass spectrometry in structural biology, biochemistry, and protein science to map and define the huge number and diversity of protein interactions * Reviews the latest discoveries and applications and addresses new and ongoing challenges This is a comprehensive reference for researchers in academia and industry engaged in studies of protein interactions and an excellent text for graduate and postgraduate students.
Author: Stephen J. Watt
Publisher:
ISBN:
Category : Electrospray ionization mass spectrometry
Languages : en
Pages : 392
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Author: M. Chance
Publisher: John Wiley & Sons
ISBN: 0470258861
Category : Science
Languages : en
Pages : 325
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Book Description
Presents a wide variety of mass spectrometry methods used to explore structural mechanisms, protein dynamics and interactions between proteins. Preliminary chapters cover mass spectrometry methods for examining proteins and are then followed by chapters devoted to presenting very practical, how-to methods in a detailed way. Includes footprinting and plistex specifically, setting this book apart from the competition.
Author: Jiangjiang Liu
Publisher:
ISBN:
Category :
Languages : en
Pages : 352
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Book Description
Since the emergence of electrospray ionization (ESI) mass spectrometry (MS) as a tool for protein structural studies, this area has experienced tremendous growth. ESI-MS is highly sensitive, and it allows the analysis of biological systems ranging in size from a few atoms to large multi-protein complexes. This work aims to solve questions in protein structural biology by using ESI-MS in conjunction with other techniques. We initially apply ESI-MS for studying the monomeric protein cytochrome c (Chapter 2). The physical reasons underlying the irreversible thermal denaturation of this protein remain controversial. By utilizing deconvoluted charge state distributions, oxidative modifications were found to be the major reason underlying the observed behavior. The positions of individual oxidation sites were identified by LC-MS/MS-based tryptic peptide mapping. Chapter 3 and 4 focus on noncovalent protein complexes. ESI allows the transfer of multi-protein complexes into the gas phase, thereby providing a simple approach for monitoring the stoichiometry of these assemblies by MS. It remains somewhat unclear, however, in how far this approach is suitable for measuring binding affinities. We demonstrate that the settings used for rf-only quadrupoles in the ion path are a key factor for ensuring uniform transmission behavior, which is a prerequisite for meaningful Kd measurements. Overall, our data support the viability of the direct ESI-MS approach for determining binding affinities of protein-protein complexes in solution. Having established suitable conditions for the analysis of noncovalent protein complexes, ESI-MS is applied for monitoring the folding and assembly of hemoglobin (Hb). The native structure of this protein comprises four heme-bound subunits. Hb represents an important model system for exploring coupled folding/binding reactions, an area that remains difficult to tackle experimentally. We demonstrate that efficient Hb refolding depends on the heme ligation status. Only under properly optimized conditions is it possible to return denatured Hb to its tetrameric native state with high yield. ESI-MS allows the observation of on-pathway and off-pathway intermediates that become populated during this highly complex self-assembly process. In summary, this work demonstrates that ESI-MS is a highly versatile tool for addressing questions at the interface of chemistry and structural biology.
Author: Amr Mostafa El-Hawiet
Publisher:
ISBN:
Category : Carbohydrates
Languages : en
Pages :
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Author: Yixuan Zhang
Publisher:
ISBN:
Category : Carbohydrates
Languages : en
Pages : 123
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Book Description
This thesis describes the development and application of electrospray ionization mass spectrometry (ESI-MS) based techniques to investigate protein-carbohydrate interactions in vitro. A catch-and-release electrospray ionization mass spectrometry (CaR-ESI-MS) assay was developed for the identification of specific interactions between water-soluble multisubunit proteins and glycosphingolipids (GSL). The assay is of high sensitivity and specificity, and demonstrates the potential for discovering biologically relevant protein-GSL interactions. Collision-induced dissociation (CID) experiments and molecular dynamic simulations were performed to investigate the dissociation pathways of multisubunit protein-ligand complexes in the gas phase. The observation of multiple dissociation pathways suggests that collisional activation of multisubunit protein-ligand complexes in the gas phase is likely to induce significant changes to the nature of the protein-ligand interactions.
Author: Julian Whitelegge
Publisher: Elsevier
ISBN: 0080932037
Category : Science
Languages : en
Pages : 563
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Book Description
This book is designed to be a central text for young graduate students interested in mass spectrometry as it relates to the study of protein structure and function as well as proteomics. It is a definite must-have work for:- libraries at academic institutions with Master and Graduate programs in biochemistry, molecular biology, structural biology and proteomics- individual laboratories with interests covering these areas - libraries and individual laboratories in the pharmaceutical and biotechnology industries. *Serves as an essential reference to those working in the field*Incorporates the contributions of prominent experts *Features comprehensive coverage and a logical structure
Author: Ling Han
Publisher:
ISBN:
Category : Electrospray ionization mass spectrometry
Languages : en
Pages : 293
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Book Description
The interactions between water-soluble proteins and carbohydrates found on the surfaces of cells play important roles in many physiological and pathological cellular processes. Carbohydrates function as receptors for signaling, cellular recognition and adhesion, and pathogen infections. This thesis focuses on the development and application of electrospray ionization mass spectrometry (ESI-MS) methods to detect and quantify protein-carbohydrate interactions in vitro. In Chapter 2, the intrinsic affinities (per binding site) of the protruding domain dimer (P dimer, 69 kDa) of the human norovirus (NoV) strain VA387 to a panel of 47 soluble analogs of histo-blood group antigens (HBGAs) were quantified using the direct ESI-MS assay. Our results revealed that the P dimer exhibits a broad specificity for the HBGAs and bind, albeit weakly (intrinsic association constants (Ka,int) of 102 - 103 M-1), to all of the oligosaccharides tested. Overall, the A and B antigens exhibit stronger binding than the H and Lewis antigens. In addition, the affinities are also affected by the precursor chain type of HBGAs but not by the chain length. In Chapter 3, the applicability of the catch-and-release (CaR)-ESI-MS assay for screening carbohydrate libraries against large protein complexes was demonstrated for the first time. Libraries containing as many as 146 compounds were screened against NoV VA387 subviral P particle (24-mer, 865 kDa). Notably, the results of the screening experiments revealed NoV interactions with oligosaccharides with structures found in human milk and the cell wall of mycobacteria. The affinities of these newly discovered ligands are comparable to those of the HBGA receptors. In Chapters 4 and 6, the direct ESI-MS assay was combined with a competitive binding strategy in order to measure the affinities of protein-carbohydrate interactions that can't be directly quantified by ESI-MS. In Chapter 4, the affinities of the NoV P particle and virus-like particle (VLP, 180-mer, 10.5 MDa) to HBGA ligands were quantified using the proxy protein ESI-MS method, which utilizes competitive protein binding. The results revealed that HBGA ligands exhibit similar affinities for the P particle and P dimer whereas the HBGA affinities for the VLP are consistently higher than those measured for the P dimer, but within a factor of three. In Chapter 6, the proxy ligand ESI-MS assay, which is on the basis of competitive ligand binding, combined with nanodisc technology to solubilize glycolipids was used to determine the interactions of cholera toxin B subunit homopentamer (CTB5) with GM1, and a family 51 carbohydrate-binding module (CBM) with B type 2 tetrasaccharide neoglycolipid. A notable finding of this study is that the affinities of the glycolipid ligands in the nanodisc are lower, by a factor of ≤5, than those of the corresponding oligosaccharides in solution. In Chapter 5, the screening using CaR-ESI-MS assay revealed the first evidence that human NoVs bind to gangliosides. Moreover, affinities measurements were reported for the NoV VA387 P dimer, P particle and VLP, and VA115 P dimer for a series of ganglioside oligosaccharides. Notably, the ganglioside affinities are similar in magnitude to those of HBGA receptors for NoVs. Additional confirmation of NoV-ganglioside interactions was provided by the binding measurements using the enzyme-linked immunosorbent assays. Finally, also in Chapter 6, a systematic ESI-MS investigation aimed at elucidating the processes that influence binding of water-soluble proteins to glycolipids incorporated into nanodiscs was described. The interactions of CTB5 to GM1 nanodiscs studied by ESI-MS indicated that proteins bind reversibly to nanodisc-associated glycolipids, and that proteins possessing multiple ligand binding sites are able to interact with glycolipids originating from different nanodiscs. Moreover, the nature of the protein-glycolipid complexes detected by ESI-MS is likely to be influenced by the diffusion of glycolipids between nanodiscs.
Author: Kevin A. Douglass
Publisher:
ISBN:
Category :
Languages : en
Pages : 190
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Book Description
Mass spectrometry is an analytical technique in which a sample is converted to gas phase ions that are subsequently separated and detected. It offers great speed, selectivity, and sensitivity during analysis, characteristics which have enabled it to become a leading method for the study of proteins. The applications of MS for these biologically significant macromolecules range from accurately determining identity and sequence to shedding light on post-translational modifications and protein molecule interactions. As a first step towards analysis by MS, gas-phase protein ions must be formed. A common method for ionization is electrospray ionization, where a liquid sample including the protein is charged, nebulized, and evaporated, resulting in bare protein ions. Although ESI has been used in this way for over two decades, many aspects of the protein charging mechanism remain unclear. To address this problem, my research has focused on identifying the factors that determine the extent of protein multiple charging during ESI and improving the ionization of proteins by desorption electrospray ionization. DESI is a method similar to ESI, except that the sample is desorbed from a surface by the spray instead of being present in it from the onset. A simple model was developed that enables the accurate prediction of protein multiple charging observed during ESI-MS if the protein sequence is known. Furthermore, the enhancement of multiple charging that is observed upon the addition of certain organic reagents, a phenomenon known as supercharging, was investigated and a novel mechanism of protein supercharging was proposed. The difficulty in analyzing large proteins by DESI-MS was studied using an innovative approach where DESI was separated into its individual sub-processes and their individual contributions to the DESI process were evaluated. As a result, core limitations to the DESI-MS of large proteins were identified. The results of my cumulative research efforts should lead to the improved MS analysis of proteins by spray ionization methods, including ESI and DESI.
