In view of rapidly growing research in the deregulation of proteases and their impact in human health and diseases, this book will highlight existing and emerging research in this exciting area. In-depth critical state-of-the-art reviews will be written by established investigators on proteases dysfunctions associated with pathogenesis of different diseases that are known to occur due to deregulation of proteolytic systems. Multidisciplinary approaches demonstrating biochemical and signal transduction mechanisms associated with deregulation of proteases leading to manifestation of the diseases will be discussed. The book highlights the roles of both intracellular and extracellular proteases in health and disease.

Membrane proteins play a key role in numerous pathologies such as cancer, cystic fibrosis, epilepsy, hyperinsulinism, and Alzheimer's disease, yet studies on these and other disorders are hampered by a lack of information about the proteins involved. In Heterologous Expression of Membrane Proteins: Methods and Protocols, expert researchers provide an overview of the different heterologous expression systems available to produce these proteins for structural analysis, helping us to understand how these proteins function, and additionally, how their functions can be modified through drug treatment. Chapters examine membrane protein solubilization, purification, and instability in solution, and investigate the strategies that helped to determine the structure of the first heterologously expressed mammalian membrane proteins. Composed in the highly successful Methods in Molecular Biology (TM) series format, each chapter contains a brief introduction, step-by-step methods, a list of necessary materials, and a Notes section which shares tips on troubleshooting and avoiding known pitfalls. Far-reaching and innovative, Heterologous Expression of Membrane Proteins: Methods and Protocols, is an essential guide for novice and experienced researchers alike, providing detailed methods for existing expression systems alongside cutting-edge strategies for future use in the field.

This second edition provides up-to-date protocols focusing on techniques for the specific enrichment of phosphopeptides and phosphoproteins. Phosphoproteomics Methods and Protocols, Second Edition guides readers through different labeling strategies for quantitative phosphoproteomics; high-throughput mass spectrometry-based phosphoproteome analyses and phospho flow cytometry; and bioinformatics strategies for phosphoproteomics data analysis and integration. Additional protocols concentrate on the identification of kinase-substrate relationships by both high- and low-throughput approaches. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Phosphoproteomics Methods and Protocols, Second Edition aims to ensure successful results in the further study of this vital field.

This volume provides a state-of-the-art update on Fc Receptors (FcRs). It is divided into five parts. Part I, Old and New FcRs, deals with the long-sought-after FcµR and the recently discovered FCRL family and TRIM21. Part II, FcR Signaling, presents a computational model of FcεRI signaling, novel calcium channels, and the lipid phosphatase SHIP1. Part III, FcR Biology, addresses major physiological functions of FcRs, their glycosylation, how they induce and regulate both adaptive immune responses and inflammation, especially in vivo, FcR humanized mice, and the multifaceted properties of FcRn. Part IV, FcRs and Disease, discusses FcR polymorphism, FcRs in rheumatoid arthritis and whether their FcRs make macaques good models for studying HIV infection. In Part V, FcRs and Therapeutic Antibodies, the roles of various FcRs, including FcγRIIB and FcαRI, in the immunotherapy of cancer and autoimmune diseases using monoclonal antibodies and IVIg are highlighted. All 18 chapters were written by respected experts in their fields, offering an invaluable reference source for scientists and clinicians interested in FcRs and how to better master antibodies for therapeutic purposes.

Calcium ions represent Mother Nature's 'ion-of-choice' for regulating fundamental physiological functions, as they initiate a new life at the time of fertilization and guide subsequent developmental and physiological functions of the human body. Calcium channels, which act as gated pathways for the movement of calcium ions across the membranes, play a central part in the initiation of calcium signals, and defects in calcium channel function have been found to result in a plethora of human diseases, referred to as the calcium channelopathies. Pathologies of Calcium Channels brings together leading international experts to discuss our current understanding of human diseases associated with the various calcium channels, from their molecular basis to potential future therapeutic targeting of calcium channels.

The field of protein NMR spectroscopy has rapidly expanded into new areas of biochemistry, molecular biology and cell biology research that were impossible to study as recently as ten years ago. This third edition of Protein NMR Techniques, expands upon the previous editions with current, detailed authoritative but down-to-earth descriptions of new methodologies. These include techniques for NMR sample preparation, solution and solid state NMR methodologies and data processing. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Protein NMR Techniques,Third Edition, seeks to aid scientists in understanding the latest innovations in the field of protein NMR.

1 a a 4 17 10 15 ubiquitin; and of 16 VCP 17 18 20 33 34 34 36 p domain. 41 42 42 43 P U 42 47 binding. C. elegans 16 In 21 22 50 51 52 53 13 and UFD 4 10 of Cdc48. 18 30 of Ufd2. COFACTORS 47 23 13 47 47 47 72 15 15 and of Spt23 p90. Ufd2 and Cdc48. In C. elegans 74 16 75 75 76 76 Ufd2 25 54 54 7 56 p47 7 7 80 30 30 81 82 82 but and CD3 26 DUB COFACTORS 30 UFD3 OTU1 4 Cdc48 30 4 OLE1. 15 27 87 REFERENCES 30 REGULATION OF UBIQUITIN MONOUBIQUITINATION UBIQUITINATION 1 32 7 S) d 33 12 13 14 15 18 19 15 20 21 35 15 15 27 15 31 32 31 33 36 monoubiquitination of pol pol 34 37 34 monoubiquitination. 20 35 trans 3 15 REFERENCES by monoubiquitination. Mol Cell; 2009. UBIQUITIN LIGASE ACTIVITY BY Nedd 1 2 of 41 5 6 8 fold. 9 13 14 edd 43 18 18 K M and k 18 22 23 K M 24 25 K M 26 edd 45 18 27 K M K D 18 25 . 8 10 M 21 28 MECHANISM AND REGULATION OF CRLs 34 41 34 edd 47 48 S. pombe 49 51 p27 and I by SCF and SCF 57 58 59 60 CTD CTD CTD CTD in Cul5 CTD CTD CTD 60 18

This second edition expands upon the previous edition with current, detailed developments in the field and brings together a multi-disciplinary team of leading researchers to provide their latest protocols for clinical proteomics analysis. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and Practical, Clinical Proteomics: Methods and Protocols, Second Edition presents approaches that will serve as a reliable guide to researchers, including clinicians, chemists, molecular biologists, bioinformaticians and computational, biologists, and investigators working on biomarker development.

Protein misfolding is a key feature of many disorders in humans, given that over twenty proteins are known to misfold and cause disease. In Protein Folding, Misfolding, and Disease: Methods and Protocols, experts in the field present a collection of current methods for studying the analysis of protein folding and misfolding, featuring strategies for expressing and refolding recombinant proteins which can then be utilized in subsequent experiments. This detailed volume also covers methods for analyzing the formation of amyloid, protocols for determining the size and structure of native and misfolded proteins, as well as specific examples of where misfolded proteins can be examined using state-of -the-art technologies. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters contain introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Up to date and authoritative, Protein Folding, Misfolding, and Disease: Methods and Protocols offers researchers the tools necessary to move ahead in this vital field.

Proteomics is a rapidly expanding investigation platform in cardiovascular medicine. Driven by major improvements in mass spectrometry (MS) instrumentation and data analysis, the proteomics field has flourished in recent years particularly in the study of complex diseases. These recent advances are characterized by the development of quantitative MS-based methods that promoted the field from primarily identifying proteins to also providing measurements of relative changes in protein levels between different cell states. This progress is reflected in the application of proteomic techniques to vascular pathology. Vascular Proteomics: Methods and Protocols provides up-to-date methods and protocols for the analysis of arteries, cells, lipoproteins, body fluids, and metabolites, with a particular focus on MS-based methods of protein and peptide quantification. Written in the successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Vascular Proteomics: Methods and Protocols is a representative selection of methods that can be a useful resource for experienced proteomics practitioners as well as newcomers interested in becoming acquainted with the practice of proteomic techniques for cardiovascular research.

Over the past decade, there has been an explosive development of research of intrinsically disordered proteins (IDPs), which are also known as unfolded proteins. Structural biologists now recognize that the functional diversity provided by disordered regions complements the functional repertoire of ordered protein regions. In Intrinsically Disordered Protein Analysis: Methods and Experimental Tools, expert researchers explore the high abundance of IDPs in various organisms, their unique structural features, numerous functions, and crucial associations with different diseases. Volume 2 includes sections on single molecule techniques, methods to assess protein size and shape, analyzing conformational behavior, mass-spectrometry, expression and purification of IDP's. Written in the highly successful Methods in Molecular Biology (TM) series format, the chapters include the kind of detailed description and implementation advice that is crucial for getting optimal results in the laboratory. Thorough and intuitive, Intrinsically Disordered Protein Analysis: Methods and Experimental Tools helps scientists further their investigations of these fascinating and dynamic molecules.

Protein analysis is increasingly becoming a cornerstone in deciphering the molecular mechanisms of life. Proteomics, the large-scale and high-sensitivity analysis of proteins, is already pivotal to the new life sciences such as Systems Biology and Systems Medicine. Proteomics, however, relies heavily on the past and future advances of protein purification and analysis methods. DIGE, being able to quantify proteins in their intact form, is one of a few methods that can facilitate this type of analysis and still provide the protein isoforms in an MS-compatible state for further identification and characterization with high analytical sensitivity. Differential Gel Electrophoresis: Methods and Protocols introduces the concept of DIGE and its advantages in quantitative protein analysis. It provides detailed protocols and important notes on the practical aspects of DIGE with both generic and specific applications in the various areas of Quantitative Proteomics. Divided into four concise sections, this detailed volume opens with the basics of DIGE, the technique and its practical details with a focus on the planning of a DIGE experiment and its data analysis. The next section introduces various DIGE methods from those employed by scientists world-wide to more novel methods, providing a glance at what is on the horizon in the DIGE world. The volume closes with an overview of the wide range of DIGE applications from Clinical Proteomics to Animal, Plant, and Microbial Proteomics applications. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters contain introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and accessible, Differential Gel Electrophoresis: Methods and Protocols can be used by novices with some background in biochemistry or molecular biology as well as by experts in Proteomics who would like to deepen their understanding of DIGE and its employment in many hyphenations and application areas. With its many protocols, applications, and methodological variants, it is also a unique reference for all who seek fundamental details on the working principle of DIGE and ideas for possible future uses of DIGE in novel analytical approaches.

In the post-genomic era, in vitro mutagenesis has emerged as a critically important tool for establishing the functions of components of the proteome. The third edition of In Vitro Mutagenesis Protocols represents a practical toolbox containing protocols vital to advancing our understanding of the connection between nucleotide sequence and sequence function. Fully updated from the previous editions, this volume contains a variety of specialty tools successfully employed to unravel the intricacies of protein-protein interaction, protein structure-function, protein regulation of biological processes, and protein activity, as well as a novel section on mutagenesis methods for unique microbes as a guide to the generalization of mutagenesis strategies for a host of microbial systems. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include brief introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and expert tips on troubleshooting and avoiding known pitfalls. Authoritative and up-to-date, In Vitro Mutagenesis Protocols, Third Edition offers today's researchers a valuable compendium of reliable and powerful techniques with which to illuminate the proteome and its rich web of biological implications.

This book describes more than 60 web-accessible computational tools for protein analysis and is totally practical, with detailed explanations on how to use these tools and interpret their results and minimal mentions to their theoretical basis (only when that is required for making a better use of them). It covers a wide range of tools for dealing with different aspects of proteins, from their sequences, to their three-dimensional structures, and the biological networks they are immersed in. The selection of tools is based on the experience of the authors that lead a protein bioinformatics facility in a large research centre, with the additional constraint that the tools should be accessible through standard web browsers without requiring the local installation of specific software, command-line tools, etc. The web tools covered include those aimed to retrieve protein information, look for similar proteins, generate pair-wise and multiple sequence alignments of protein sequences, work with protein domains and motifs, study the phylogeny of a family of proteins, retrieve, manipulate and visualize protein three-dimensional structures, predict protein structural features as well as whole three-dimensional structures, extract biological information from protein structures, summarize large protein sets, study protein interaction and metabolic networks, etc. The book is associated to a dynamic web site that will reflect changes in the web addresses of the tools, updates of these, etc. It also contains QR codes that can be scanned with any device to direct its browser to the tool web site. This monograph will be most valuable for researchers in experimental labs without specific knowledge on bioinformatics or computing.

Modification of target protein properties by reversible phosphorylation events has been found to be one of the most prominent cellular control processes in all organisms. Recent advances in the areas of molecular biology and biochemistry are presenting new possibilities for reaching an unprecedented depth and a proteome-wide understanding of phosphorylation processes in plants as well as in other species. The major goal of Plant Kinases: Methods and Protocols is to provide the experimentalist with a detailed account of the practical steps necessary for successfully carrying out each protocol in his or her own laboratory. Plant protein kinases specifically addressed in this volume are members of the plant MAP kinase cascade, cyclin- and Calcium-dependent protein kinases, and plant sensor and receptor kinases. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters contain introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and accessible, Plant Kinases: Methods and Protocols will prove a useful laboratory companion to both novice and seasoned researchers by facilitating the practical work that will lead them to new and exciting insights in this dynamic field.

Featuring a diverse array of model organisms and scientific techniques, Sirtuins: Methods and Protocols collects detailed contributions from experts in the field addressing this vital family of genes. Opening with methods to generate sirtuin biology tools, the book continues by covering methods to identify sirtuin substrates, to measure sirtuin activity, and to study sirtuin biology. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Comprehensive and easy to use, Sirtuins: Methods and Protocols presents detailed protocols for sirtuin research that can be followed directly or modified to investigate new areas of sirtuin biology.

Glycosylation is the most abundant post-translational modification of proteins. Estimates vary widely, but a common assessment is that upwards of 50% of eukaryotic proteins are modified by some type of glycan. In Mass Spectrometry of Glycoproteins: Methods and Protocols, expert researchers in the field detail many of the methods that are now commonly used for glycoproteomics. These methods and techniques include robust sample preparation techniques; advanced chromatographic strategies for improving dynamic range; state-of-the-art mass spectrometry instrumentation and associated ionization and fragmentation methods; and informatics tools used for identifying glycoproteins and characterizing the associated glycans. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Mass Spectrometry of Glycoproteins: Methods and Protocol is an essential resource for those who work at the interface of glycobiology and mass spectrometry.

Since the second half of the 20th century machine computations have played a critical role in science and engineering. Computer-based techniques have become especially important in molecular biology, since they often represent the only viable way to gain insights into the behavior of a biological system as a whole. The complexity of biological systems, which usually needs to be analyzed on different time- and size-scales and with different levels of accuracy, requires the application of different approaches, ranging from comparative analysis of sequences and structural databases, to the analysis of networks of interdependence between cell components and processes, through coarse-grained modeling to atomically detailed simulations, and finally to molecular quantum mechanics. This book provides a comprehensive overview of modern computer-based techniques for computing the structure, properties and dynamics of biomolecules and biomolecular processes. The twenty-two chapters, written by scientists from all over the world, address the theory and practice of computer simulation techniques in the study of biological phenomena. The chapters are grouped into four thematic sections dealing with the following topics: the methodology of molecular simulations; applications of molecular simulations; bioinformatics methods and use of experimental information in molecular simulations; and selected applications of molecular quantum mechanics. The book includes an introductory chapter written by Harold A. Scheraga, one of the true pioneers in simulation studies of biomacromolecules.

Pulse Dipolar Electron Spin Resonance: Distance Measurements by Peter P. Borbat, Jack H. Freed.Interpretation of Dipolar EPR Data in Terms of Protein Structure, by Gunnar Jeschke.Site-Directed Nitroxide Spin Labeling of Biopolymers, by Sandip A. Shelke and Snorri Th. Sigurdsson. Metal-Based Spin Labeling for Distance Determination, by Daniella Goldfarb. Structural Information from Spin-Labelled Membrane-Bound Proteins, by Johann P. KLare, Heinz-Jürgen Steinhoff. Structural Information from Oligonucleotides, by Richard Ward and Olav Schiemann. Orientation selective DEER using rigid spin labels, cofactors, metals, and clusters, by Claudia E. Tait, Alice M. Bowen, Christiane R. Timmel, Jeffrey Harmer

Transgenic Plants: A Production System for Industrial and Pharmaceutical Proteins provides a detailed guide to the principles and practice of using transgenic plants as a system for the production of heterologous proteins. It is unique in that it covers the complete process of heterologous protein production in plants, from the initial transformation of the plant, through to transcription, transgene stability and finally the downstreaming processing events for protein purification. Written by an international team of industrialists and academics, this book describes:

• the fundamental issues associated with expressing heterologous proteins in plants;
• a number of detailed examples of the successful small-and large-scale production of proteins;
• the essentials of patenting; and
• the commercial exploitation.

Lectins: Methods and Protocols summarizes classic lectin technologies and advanced techniques with high throughputs and sensitivities. Chapters include methods and techniques for serial lectin-affinity chromatography procedure, lectin-probed western blot and histochemical analyses, quantitative interaction analyses based on equilibrium dialysis, isothermal calorimetry, surface plasmon resonance, techniques for elucidating functions of endogenous animal lectins, advanced methods on "engineer" novel lectins by evolutionary concepts, and special equipment. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Lectins: Methods and Protocols seeks to aid not only lectin specialists but also non-experts including both young scientists.

In a simplified form, epigenetics refers to heritable changes in phenotype that are not due to changes in the underlying DNA sequence. In this book, epigenetic mechanisms of regulation and dysregulation in health and disease are explored in great depth. Detailed chapters on epigenetic processes including DNA methylation and chromatin post-translational modifications including potential interventions with DNA methyltransferase inhibitors and histone deacetylase inhibitors are explored in initial chapters. These provide a detailed overview and important background to the entire field. The book is then focussed on epigenetic mechanisms involved in various diseases including anti-inflammatory and autoimmune conditions. Important accounts relating to the effects of epigenetics in metabolic syndrome, cardiovascular disease and asthma are the focus of subsequent chapters. The role of epigenetic dysregulation in malignancy is a current topic of interest and represents an intense field of research. A large component of this book is dedicated to the analysis of aberrant epigenetic processes in carcinogenesis and cancer progression. Further, chapters are focused on emerging cancer prevention using nutritional components and anti-cancer therapies particularly with histone deacetylase inhibitors, which have already been approved for the treatment of cutaneous T-cell lymphoma. The emerging role of nanoparticle preparations, especially in the context of delivering potential epigenetic therapies to target cells in various diseases, is also explored in this book. Overall, this book encompasses a wide range of topics related to epigenetic mechanisms in health and disease and would appeal to anyone with an interest in epigenetics, chromatin biology and emerging epigenetic interventions and therapies.

In her thesis, Sara Bobone outlines spectroscopic studies of antimicrobial peptides (AMPs) which are promising lead compounds for drugs used to fight multidrug resistant bacteria. Bobone shows that AMPs interact with liposomes and she clarifies the structure of pores formed by one of these molecules. These results help us to understand how AMPs are selective for bacterial membranes and how their activity can be finely tuned by modifying their sequence. Findings which solve several conundrums debated in the literature for years. In addition, Bobone uses liposomes as nanotemplates for the photopolymerization of hydrogels - exploiting the self- assembly properties of phospholipids. Bobone was able to trap an enzyme using nanometeric particles, while still allowing its activity by the diffusion of substrates and products through the network of the polymer. The innovative nano devices described in this thesis could solve many of the hurdles still hampering the therapeutic application of protein-based drugs.

This volume describes our current understanding of the biological role of visual and non-visual arrestins in different cells and tissues, focusing on the mechanisms of arrestin-mediated regulation of GPCRs and non-receptor signaling proteins in health and disease. The book covers wide range of arrestin functions, emphasizing therapeutic potential of targeting arrestin interactions with individual partners.

Galectins: Methods and Protocols is the first book solely dedicated to methodological approaches designed to study galectin function. The galectin family represents one of the most pleiotropic families, with individual members having been implicated in various aspects of nearly every biological process described, from RNA splicing to complex regulatory circuits that orchestrate adaptive immunity. Given the diverse roles of galectins in a variety of biological systems, studying these glycan binding proteins often requires the assimilation of diverse technical skills to fully appreciate their biological function. Their nearly ubiquitous expression and ability to bind highly modifiable carbohydrate ligands, in addition to a variety of other regulatory proteins, allows these glycan binding proteins (GBPs) to possess the capacity to regulate a wide variety of biological processes. Individual chapters are dedicated to examining salient features of galectin functions. Written in the successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Galectins: Methods and Protocols seeks to serve both professionals and novices with a useful framework when examining galectin function for many years to come.