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.

Beginning from centuries of anecdotal descriptions of cell death, such as those on the development of the midwife toad in 1842 by Carl Vogt, to modern-day investigations of cell death as a biological discipline, it has become accepted that cell death in multicellular organisms is a normal part of life. This book provides a comprehensive view of cell death, from its mechanisms of initiation and execution, to its implication in human disease and therapy. Physiological cell death plays critical roles in almost all aspects of biology, and the book details its roles in lymphocyte homeostasis, neuronal function, metabolism, and the DNA damage response. When physiological cell death goes awry, diseases can arise, and cancer is presented as a central paradigm for the consequences of derangements in the interplay between cell survival and cell death. At the same time, the potential promise of targeted therapies aimed at interdicting cell death machineries are also discussed extensively. The molecular mechanisms that underlie apoptotic cell death are illustrated from the perspectives of both the intrinsic, mitochondrial apoptotic pathway and the extrinsic, death receptor pathway. Key players in these pathways, such as the Bcl2 family proteins, cytochrome c, Apaf-1, caspases, death receptor adapter proteins, and inhibitor of apoptosis proteins, are presented from both functional and structural angles. Until only a few years ago, programmed cell death has been considered essentially synonymous with apoptosis. However, we now know that programmed cell death can also take other forms such as necrosis or necroptosis, and to this end, the mechanisms that underlie programmed necrosis in development and host defense are illustrated. The past twenty plus years have seen an incredible growth of research in cell death, with one breakthrough after another, and the legacy still goes on with constant new surprises and findings. Long live cell death!

In Fluorescent Protein-Based Biosensors: Methods and Protocols, experts in the field have assembled a series of protocols describing several methods in which fluorescent protein-based reporters can be used to gain unique insights into the regulation of cellular signal transduction. Genetically encodable fluorescent biosensors have allowed researchers to observe biochemical processes within the endogenous cellular environment with unprecedented spatiotemporal resolution. As the number and diversity of available biosensors grows, it is increasingly important to equip researchers with an understanding of the key concepts underlying the design and application of genetically encodable fluorescent biosensors to live cell imaging. 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, Fluorescent Protein-Based Biosensors: Methods and Protocols promises to be a valuable resource for researchers interested in applying current biosensors to the study of biochemical processes in living cells as well as those interested in developing novel biosensors to visualize other cellular phenomena.

This book provides a molecular view of membrane transport by means of numerous biochemical and biophysical techniques. The rapidly growing numbers of atomic structures of transporters in different conformations and the constant progress in bioinformatics have recently added deeper insights. The unifying mechanism of energized solute transport across membranes is assumed to consist of the conformational cycling of a carrier protein to provide access to substrate binding sites from either side of a cellular membrane. Due to the central role of active membrane transport there is considerable interest in deciphering the principles of one of the most fundamental processes in nature: the alternating access mechanism. This book brings together particularly significant structure-function studies on a variety of carrier systems from different transporter families: Glutamate symporters, LeuT-like fold transporters, MFS transporters and SMR (RND) exporters, as well as ABC-type importers. The selected examples impressively demonstrate how the combination of functional analysis, crystallography, investigation of dynamics and computational studies has made it possible to create a conclusive picture or more precisely, "a molecular movie". Although we are still far from a complete molecular description of the alternating access mechanism, remarkable progress has been made from static snapshots towards membrane transport dynamics.

Defining and understanding cellular and molecular mechanisms that are relevant to women's health has become a critical area of scientific pursuit. Until recently, very little effort has been place on defining or understanding critical differences between women and men that may be critical to the overall health of the woman. In 1990, the National Institutes of Health recognized this gap in knowledge resulting in the creation of the Office of Research on Women's Health. One of the purposes of this office was to advance the understanding of health issues from the women's perspective from both a basic and clinical scientific perspective. From a scientific evolution of understanding, the existence of this office is new and thus there has not been enough time for new information to integrate itself in our current scientific thought process. This book will seek to capture and disseminate our current understanding of scientific advancements relevant to women's health and provide the information to a broad audience. The purpose of this work is to discuss recent advancements in basic science across three areas of concern for women's health. In addition, the book will provide "translational" chapters that attempt to place the basic science work in context within our current understanding of the human. Although it is well acknowledge that gender differences exist across organ function which translates into differences in whole body function, until recently little effort has been made to define basic mechanisms within various tissues within the woman. This work will focus on recent scientific findings that are relevant to women's health and to provide novel and relevant information to interested scientists and clinicians.

Remarkably, while G protein-coupled receptors (GPCRs) are highly prevalent in animals and yeast, very few candidate GPCRs have been identified in plants. In G Protein-Coupled Receptor Signaling in Plants: Methods and Protocols, experts in the field describe techniques used in the study of small GTPases and related proteins. Beginning with a chapter on bioinformatics approaches for GPCR discovery, this detailed volume continues with chapters on heterotrimeric G protein subunits, Rab-GTPases, as well as lipid modifications, including myristoylation, acylation, and prenylation. 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. Practical and dependable, G Protein-Coupled Receptor Signaling in Plants: Methods and Protocols aims to aid further studies into the roles of small GTPases which will help elucidate numerous key processes in plants.

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.

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.

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.

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.

The present book gives an overview on the similarities and differences of the various translation systems. Moreover, it highlights the mechanisms and control of translation in mitochondria and other organelles such as chloroplasts, plastids and apicoplasts in different organisms. Lastly, it offers an outlook on future developments and applications that might be made possible by a better understanding of translation in mitochondria and other organelles.  

Since the publishing of the first edition, the methodologies and instrumentation involved in the field of mass spectrometry-based proteomics has improved considerably. Fully revised and expanded, Mass Spectrometry Data Analysis in Proteomics, Second Edition presents expert chapters on specific MS-based methods or data analysis strategies in proteomics. The volume covers data analysis topics relevant for quantitative proteomics, post translational modification, HX-MS, glycomics, and data exchange standards, among other topics. Written in the highly successful Methods in Molecular Biology series format, chapters include brief introductions to their respective subjects, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Updated and authoritative, Mass Spectrometry Data Analysis in Proteomics, Second Edition serves as a detailed guide for all researchers seeking to further our knowledge in the field of proteomics.

This book systematically summarizes the ideas and technologies used in urine proteome analysis. It argues that change is the core of biomarker definition since the body uses its homeostatic mechanisms to correct changes in the blood. This means that urine is probably a better source of biomarkers than blood. A roadmap to the urinary biomarker era is proposed, and researchers are reminded of the potential opportunities and risks in their study design. Kidney diseases are emphasized as they produce the most significant changes in urine. This book tries to show researchers and graduate students, who are in or entering the field, "all things considered" rather than "the current affair".

Focusing on model systems for the study of structure, folding, and association in the membrane, Membrane Proteins: Folding, Association, and Design presents an overview of methods that can be applied to these intricate systems. The volume is divided into four detailed sections, covering association of transmembrane helices, interactions with the lipid bilayer, NMR methods, as well as a variety of engineering approaches. Written for the highly successful Methods in Molecular Biology series, 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, Membrane Proteins: Folding, Association, and Design serves as an ideal guide for researchers reaching for the tantalizing possibility of designing novel membrane proteins with tailored functionality.

Protein Structure Prediction, Third Edition expands on previous editions by focusing on software and web servers. With new chapters that provide instructions on how to use a computational method with examples of prediction by the method. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials, step-by-step, readily reproducible protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Protein Structure Prediction, Third Edition provides diverse methods detailing the expansion of the computational protein structure prediction field.

This brief discusses the mechanism of functional expression of a protein or protein complex utilizing the ATP hydrolysis cycle or proton-motive force from a unique point of view focused on the roles of water. A variety of processes are considered such as the unidirectional movement of a linear-motor protein along a filament, insertion of an unfolded protein into a chaperonin and release of the folded protein from it, transport of diverse substrates across the membrane by a transporter, and directed rotation of the central subunit within a rotatory motor protein complex. These topics are discussed in a unified manner within the same theoretical framework. The author argues that water plays imperative roles in the functional expression of these molecular machines. A pivotal factor is the entropic force or potential originating from the translational displacement of water molecules coexisting with the molecular machines in the entire system.

Presents Practical Applications of Mass Spectrometry for Protein Analysis and Covers Their Impact on Accelerating Drug Discovery and Development * Covers both qualitative and quantitative aspects of Mass Spectrometry protein analysis in drug discovery * Principles, Instrumentation, Technologies topics include MS of peptides, proteins, and ADCs , instrumentation in protein analysis, nanospray technology in MS protein analysis, and automation in MS protein analysis * Details emerging areas from drug monitoring to patient care such as Identification and validation of biomarkers for cancer, targeted MS approaches for biomarker validation, biomarker discovery, and regulatory perspectives * Brings together the most current advances in the mass spectrometry technology and related method in protein analysis

This Brief explores the use of proteomics as a tool for biomarker discovery in human reproduction and summarizes current findings and trends of proteomic studies in both male and female infertility. This simplifies this important but complex topic and equips the novice reader with sufficient background information on the use of proteomics in human reproduction. The up-to-date scenario on proteomic investigations will also appeal to researchers and post graduate students looking to keep abreast with the latest developments in reproductive research. This review summarizes current findings of contemporary proteomic studies on infertility in both males and females with various reproductive pathologies, and its use in predicting the outcome of assisted reproduction. In human reproduction, the search for biomarkers via proteomics is a fast-evolving approach that involves the analysis of proteins in the reproductive tissues and fluids, such as the male gametes, seminal plasma, ovarian and endometrial tissue, and follicular and uterine fluid. By comparing the protein profile of a healthy, fertile individual against that of an infertile individual, the differentially expressed proteins may give an indication to certain proteins that could serve as useful biomarkers that are related to infertility. As proteomic studies continue to unravel the dynamic proteome behind various infertility conditions, there is potential for the discovery of prognostic markers that could ultimately help in both natural and assisted human reproduction.

This detailed book expands upon the previous edition with a collection of methods for those performing experimental work on small GTPases of the Rho family. Split into four sections, the volume explores computational modeling and imaging procedures, biochemical methods related to post-translational modifications of Rho GTPases as well as some high throughput methods, functional assays that allow for monitoring the consequences of manipulating Rho GTPases in a variety of cell types and cell biology processes, and techniques specifically designed for studies in selected non-mammalian model organisms (zebrafish, social ameba, plants and algae). Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on trouble shooting and avoiding known pitfalls. Authoritative and up-to-date, Rho GTPases: Methods and Protocols, Second Edition constitutes an invaluable tool for all those with an interest in this remarkable family of signaling proteins.

G protein-coupled receptors (GPCRs) are heptahelical transmembrane receptors that convert extra-cellular stimuli into intra-cellular signaling, and ultimately into biological responses. Since GPCRs are natural targets for approximately 40% of all modern medicines, it is not surprising that they have been the subject of intense research. Notwithstanding the amount of data generated over the years, discovering ligands of these receptors with optimal therapeutic properties is not straightforward and has certainly been hampered for years by the lack of high-resolution structural information about these receptors. Luckily, there has been a steady increase of high-resolution crystal structures of these receptors since 2007, and this information, integrated with dynamic inferences from computational and experimental methods, holds great potential for the discovery of new, improved drugs. This book, which provides, for the first time, state-of-the-art views on modeling and simulation of GPCRs, is divided into 4 parts. In the first part, the impact of currently available GPCR crystal structures on structural modeling is discussed extensively as are critical insights from simulations in the second part of the book. The third part reports recent progress in rational ligand discovery and mathematical modeling, whereas the fourth part provides an overview of bioinformatics tools and resources that are available for GPCRs.

This book addresses the differentiation control of skeletal muscle in different locations of the vertebrate body Particular attention is paid to novel regulatory molecules and signals as well as the heterogeneity of origin that have revealed a developmental overlap between skeletal and cardiac muscle. Different functional muscle groups are the product of the evolution of the vertebrate classes, making a phylogenetic comparison worthwhile for understanding the role of muscle stem cells and precursors in myogenesis. New insights into the hierarchy of transcription factors, particularly in the context of these different muscle groups have been gained from detailed investigations of the spatio-temporal and regulatory relationships derived from mouse and zebrafish genetics and avian microsurgery. Importantly, epigenetic mechanisms that have surfaced recently, in particular the role of MyomiRs, are also surveyed. With an eye to the human patient, encouraging results have been generated that identify parallels between embryonic myogenesis and regenerating myofibers due to common regulatory molecules. On the other hand, both processes differ considerably in quality and complexity of the processes employed. Interestingly, the heterogeneity in embryonic sources from which skeletal muscle groups in the vertebrate including the human body take origin is paralleled by differences in their susceptibility to particular muscle dystrophies as well as by the characteristics of the satellite cells involved in regeneration. The progress that has been made in the field of muscle stem cell biology, with special focus on the satellite cells, is outlined in this book by experts in the field. The authors review recent insights of the heterogeneous nature of these satellite cells regarding their gene signatures and regeneration potential. Furthermore, an improved understanding of muscle stem cells seems only possible when we study the impact of the cell environment on efficient stem cell replacement therapies for muscular dystrophies, putting embryological findings from different vertebrate classes and stem cell approaches into context.

With insolubility proving to be one of the most crippling bottlenecks in the protein production and purification process, this volume serves to aid researchers working in the recombinant protein production field by describing a wide number of protocols and examples. Insoluble Proteins: Methods and Protocols includes chapters that describe not only the recombinant protein production in different expression systems but also different purification and characterization methods to finally obtain these difficult-to-obtain proteins. Beginning with protein production methods using both prokaryotic and eukaryotic expression systems, the book continues with purification protocols using insoluble proteins, the characterization of insoluble proteins, as well as a general overview of interesting applications of insoluble proteins. 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 practical, Insoluble Proteins: Methods and Protocols aims to provide the scientific community with detailed and reliable state-of-the-art protocols that are used in order to successfully produce and purify recombinant proteins prone to aggregate.

Exploring these type II trans-membrane proteins, The TNF Superfamily: Methods and Protocols focuses on various techniques to investigate aspects of the TNF Superfamily members in health and disease. Opening with protocols to understand the signaling process of TNF family members, this detailed volume continues with technical examples of investigating the role of TNF family members in physiopathologies, protocols on modulation of TNF signaling by pathogens, experimental applications of TNF-reporter mice, as well as methodologies for various assays of TNF family members and the production of recombinant molecules. Written for the Methods in Molecular Biology series, 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. Practical and ready to use, The TNF Superfamily: Methods and Protocols will aid researchers investigating this key family of proteins, involved in vital processes such as providing signals for activation, differentiation, survival and death of cells, modulation of immune response and inflammation, hematopoiesis and osteoclastogenesis.

NMR spectroscopy has proven to be a powerful technique to study the structure and dynamics of biological macromolecules. Fundamentals of Protein NMR Spectroscopy is a comprehensive textbook that guides the reader from a basic understanding of the phenomenological properties of magnetic resonance to the application and interpretation of modern multi-dimensional NMR experiments on 15N/13C-labeled proteins. Beginning with elementary quantum mechanics, a set of practical rules is presented and used to describe many commonly employed multi-dimensional, multi-nuclear NMR pulse sequences. A modular analysis of NMR pulse sequence building blocks also provides a basis for understanding and developing novel pulse programs. This text not only covers topics from chemical shift assignment to protein structure refinement, as well as the analysis of protein dynamics and chemical kinetics, but also provides a practical guide to many aspects of modern spectrometer hardware, sample preparation, experimental set-up, and data processing. End of chapter exercises are included to emphasize important concepts. Fundamentals of Protein NMR Spectroscopy not only offer students a systematic, in-depth, understanding of modern NMR spectroscopy and its application to biomolecular systems, but will also be a useful reference for the experienced investigator.

Each review within the volume critically surveys one aspect of that topic and places it within the context of the volume as a whole. The most significant developments of the last 5 to 10 years are presented using selected examples to illustrate the principles discussed. The coverage is not intended to be an exhaustive summary of the field or include large quantities of data, but should rather be conceptual, concentrating on the methodological thinking that will allow the non-specialist reader to understand the information presented. Contributions also offer an outlook on potential future developments in the field.