This volume explores strategies and detailed protocols for the preparation of macromolecular complexes and their characterization in view of structural analysis. The chapters in this book are separated into three parts: Part One focuses on sample preparation, and covers strategies for recombinant expression of multiprotein complexes in prokaryotic and eukaryotic hosts, for genome engineering using the CRISPR/Cas9 system and for production of specific binders such as reformatted antibodies and artificial binding proteins. Part Two looks at the biophysical methods that can provide useful indicators for sample optimization, and often complement structural information obtained with core technologies for structure determination-x-ray crystallography and cryo-electron microscopy-by quantitative solution data. Part Three discusses the characterization of multiprotein complexes in a cellular environment using the latest technologies and in vivo 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, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Cutting-edge and authoritative, Multiprotein Complexes: Methods and Protocols is a valuable resource for structural and molecular biologists who need to prepare multi-components for their applications, and for other scientists working on macromolecular assemblies from other angles that need to know the latest approaches that the field has to offer.

The study of carbonic anhydrase has spanned multiple generations of scientists. Carbonic anhydrase was first discovered in 1932 by Meldrum and Roughton. Inhibition by sulfanilamide was shown in 1940 by Mann and Keilin. Even Hans Krebs contributed to early studies with a paper in 1948 showing the relationship of 25 different sulfonamides to CA inhibition. It was he who pointed out the importance of both the charged and uncharged character of these compounds for physiological experiments. The field of study that focuses on carbonic anhydrase (CA) has exploded in recent years with the identification of new families and isoforms. The CAs are metalloenzymes which are comprised of 5 structurally different families: the alpha, beta, gamma, and delta, and epsilon classes. The alpha class is found primarily in animals with several isoforms associated with human disease. The beta CAs are expressed primarily in plants and are the most divergent. The gamma CAs are the most ancient. These are structurally related to the beta CAs, but have a mechanism more similar to the alpha CAs. The delta CAs are found in marine algae and diflagellates. The epsilon class is found in prokaryotes in which it is part of the carboxysome shell perhaps supplying RuBisCO with CO2 for carbon fixation. With the excitement surrounding the discovery of disease-related CAs, scientists have redoubled their efforts to better understand structure-function relationships, to design high affinity, isotype-specific inhibitors, and to delineate signaling systems that play regulatory roles over expression and activity. We have designed the book to cover basic information of mechanism, structure, and function of the CA families. The authors included in this book bring to light the newest data with regard to the role of CA in physiology and pathology, across phylums, and in unique environmental niches.

Main Question: G protein coupled receptors are involved in highly efficient and specific activation of signalling pathways. How do GPCR signalling complexes get assembled to generate such specificity? In order to answer this question, we need to understand how receptors and their signalling partners are synthesized, folded and quality-controlled in order to generate functional proteins. Then, we need to understand how each partner of the signalling complex is selected to join a complex, and what makes this assembly possible. GPCRs are known to be able to function as oligomers, what drives the assembly into oligomers and what will be the effects of such organization on specificity and efficacy of signal transduction. Once the receptor complexes are assembled, they need to reach different locations in the cell; what drives and controls the trafficking of GPCR signalling complexes. Finally, defects in synthesis, maturation or trafficking can alter functionality of GPCRs signalling complexes; how can we manipulate the system to make it function normally again? Pharmacological chaperones may just be part of the answer to this question.

This is the second edition of a very well received book that details how the sumoylation system functions and how it modulates numerous cellular activities. SUMO is a post-translational modifier in the ubiquitin super-family that has gained recognition over the last twenty years as an essential and prevalent regulatory molecule. Individual chapters explore the biochemistry, molecular biology, and cell biology of the sumoylation system and its substrate proteins. The book is divided into three themed parts: Molecular Functions (I), Cell Growth Regulation (II), and Diseases (III). Parts I and II focus on the contribution of sumoylation to cellular activities in both the nuclear and cytoplasmic compartments. The nuclear activities covered include nucleic acid metabolism (both RNA and DNA), chromosome structure and replication, and nucleocytoplasmic transport. Cytoplasmic processes presented include regulation of membrane ion channels, general metabolism, and apoptotic signalling. Topics in Part III include the role of sumoylation in developmental abnormalities (craniofacial and cardiovascular), diabetes, neurodegenerative diseases, cancer, and infections with viruses and bacteria. Each of the corresponding chapter authors is an active researcher who has made significant contributions to understanding sumoylation. This second edition provides updates and revisions to most of the original chapters plus adds six new chapters to address important developing areas of sumoylation research. This volume is intended for a scientific audience from undergraduates to independent researchers. The content will serve as both a solid introduction for the novice reader and an in depth treatment for the advanced scholar.

This second edition provides new and updated methods on the principles underlying modern protein analysis, from statistical issues to gel-based and mass spectrometry-based applications. Chapters detail protein quantification as basis for realisation of quantitative studies, gel-based and mass spectrometry-based quantification techniques, TMT, IPTL, PRM, MALDI Imaging, SILAC, PTM analysis, DIA, cross-linking, and the up-to-date topics of software and data 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 cutting-edge, Quantitative Methods in Proteomics, Second Edition aims to provide comprehensive and competent overview in the important and still growing field of quantitative proteomics.

A NATO Advanced Research Workshop entitled New Methods for the Study of Molecular Aggregates was held at Tbe Lodge at Kananaskis Village, Alberta, Canada from 16 -20 June 1996. In fact the meeting was entirely concerned with the problem of analyzing biomolecular complexes, so the title of these proceedings has been altered to give a more precise description of the content. Tbe workshop was hosted by the time-of-flight group of the Department of Physics at the University of Manitoba, and was attended by 64 participants from around the world. '!\venty-one invited talks were given and 27 papers were presented as posters. Of the 48 contributions, 22 papers (12 orals, 10 posters) are included in these proceedings. Tbe subject of the conference was the investigation of noncovalent biomolecular complexes, with particular focus on the application of mass spectrometry to their characterization. '!\vo new ionization techniques introduced in the late 1980s, electrospray ionization (ES I) and matrix-assisted laser desorptionlionization (MALDI), resulted in a breakthrough in mass spectrometry, enabling its use in molecular weight and primary structure determination of biopolymers larger than 100 kDa. Recently it has been discovered that ESI mass spectrometry mayaiso be used to characterize complexes containing noncovalent interactions, thus opening new perspectives for supramolecular chemistry. ESI mass spectrometry has the advantage that the sampie is introduced from a homogenous solution which can be maintained at near physiological conditions of pR, concentration, and temperature.

First published in 1943, Vitamins and Hormones is the longest-running serial published by Academic Press. The Series provides up-to-date information on vitamin and hormone research spanning data from molecular biology to the clinic. A volume can focus on a single molecule or on a disease that is related to vitamins or hormones. A hormone is interpreted broadly so that related substances, such as transmitters, cytokines, growth factors and others can be reviewed. This volume focuses on nociceptin opioid.

The need for information in the understanding of membrane systems has been caused by three things - an increase in computer power; methodological developments and the recent expansion in the number of researchers working on it worldwide. However, there has been no up-to-date book that covers the application of simulation methods to membrane systems directly and this book fills an important void in the market. It provides a much needed update on the current methods and applications as well as highlighting recent advances in the way computer simulation can be applied to the field of membranes and membrane proteins. The objectives are to show how simulation methods can provide an important contribution to the understanding of these systems. The scope of the book is such that it covers simulation of membranes and membrane proteins, but also covers the more recent methodological developments such as coarse-grained molecular dynamics and multiscale approaches in systems biology. Applications embrace a range of biological processes including ion channel and transport proteins. The book is wide ranging with broad coverage and a strong coupling to experimental results wherever possible, including colour illustrations to highlight particular aspects of molecular structure. With an internationally respected list of authors, its publication is timely and it will prove indispensable to a large scientific readership.

New insights into modern medicine and systems biology are enabled by innovative protocols and advanced technologies in mass spectrometry-based proteomics. This volume details new pipelines, workflows, and ways to process data that allow for new frontiers in proteomics to be pushed forward. With applications to biomarker discovery, interactions between proteins, between biological systems, dynamics of post-translational modifications among others, new protocols have been developed and iteratively refined to probe the endless complexity of the proteome in ever greater details. This volume deals with methods for data dependent and data independent mass spectrometry analyses. Valuable, first-hand information is provided from designing experiments, sample preparation and analysis, exploitation of public datasets and carrying out reproducible data pipelines, using modern computational tools such as Galaxy or Jupyter. 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 cutting-edge, Mass Spectrometry of Proteins: Methods and Protocols aims to ensure successful results in the further study of this vital field.

A Unified Microscopic Approach to Analyzing Complex Processes in Molecular Motors Motor Proteins and Molecular Motors explores the mechanisms of cellular functioning associated with several specific enzymatic molecules called motor proteins. Motor proteins, also known as molecular motors, play important roles in living systems by supporting cellular transport and force generation via the transformation of chemical energy into mechanical work. The book presents established results, theoretical methods, and experimental observations related to biological molecular motors. It uses fundamental physical-chemical concepts and methods to develop a systematic theoretical framework for understanding motor protein dynamics. The author introduces the main ideas using simple arguments that avoid heavy mathematical derivations in favor of more intuitive physical understanding. Although the book assumes some rudimentary knowledge of cell biology, calculus, and basic ideas from chemistry and physics, it gives explanations and derivations for most results. Accessible to students and researchers in a wide range of scientific fields, this book provides a unified molecular picture for analyzing motor proteins. It connects major experimental facts on molecular motors to principal theoretical concepts consistent with the fundamental laws of chemistry and physics.

The edition details methods to study intrinsically disordered proteins (IDPs) including recent topics such as extremely high-affinity disordered complexes, kinetics that evade established concepts, liquid-liquid phase separation, and novel disorder-driven allosteric mechanisms. 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 cutting-edge, Intrinsically Disordered Proteins: Methods and Protocols aims to help scientists with different backgrounds to further their investigations into these fascinating and dynamic molecules. Chapter 24 is available open access under a CC BY 4.0 license via link.springer.com. Chapters "40 and 42 " are available open access under a Creative Commons Attribution 4.0 International License via link.springer.com.

Daniel C. Liebler masterfully introduces the science of proteomics by spelling out the basics of how one analyzes proteins and proteomes, and just how these approaches are then employed to investigate their roles in living systems. He explains the key concepts of proteomics, how the analytical instrumentation works, what data mining and other software tools do, and how these tools can be integrated to study proteomes. Also discussed are how protein and peptide separation techniques are applied in proteomics, how mass spectrometry is used to identify proteins, and how data analysis software enables protein identification and the mapping of modifications. In addition, there are proteomic approaches for analyzing differential protein expression, characterizing proteomic diversity, and dissecting protein-protein interactions and networks.

This book is the first example in presenting LC-MS strategies for the analysis of peptides and proteins with detailed information and hints about the needs and problems described from experts on-the-job. The best advantage is -for sure- the practical insight of experienced analysts into their novel protein analysis techniques. Readers starting in 'Proteomics' should be able to repeat each experiment with own equipment and own protein samples, like clean-up, direct protein analysis, after (online) digest, with modifications and others. Furthermore, the reader will learn more about strategies in protein analysis, like quantitative analysis, industrial standards, functional analysis and more.

Protein phosphorylation analysis is a central theme in current analytical biochemistry, cell biology and systems biology. Due to its versatility, specificity and sensitivity, mass spectrometry has developed into a key technology in this field. A set of minor and major instrumental innovations mean that mass spectrometers now exhibit a level of performance, a stability of operation, a relative ease of use, and productivity, which would once have been hard to imagine. This book guides the reader through this prolific field by presenting a collection of personal views and selected examples which cover all the important principles with a focus on electrospray ionization mass spectrometry. It covers: phosphorylation analysis at the peptide, protein and proteome level; manual and automated data evaluation; phosphopeptide enrichment; quantitative aspects; element mass spectrometry; individual analytical strategies, and hints to useful internet resources. This book provides students, graduate students, post-Docs and senior scientists from related areas with a better understanding on molecular protein phosphorylation analysis. Its highest aim is to strengthen the reader's ability to develop a personal, well-founded opinion on original manuscripts published in this field.

Introduction to Computational Proteomics introduces the field of computational biology through a focused approach that tackles the different steps and problems involved with protein analysis, classification, and meta-organization. The book starts with the analysis of individual entities and works its way through the analysis of more complex entities, from protein families to interactions, cellular pathways, and gene networks. The first part of the book presents methods for identifying the building blocks of the protein space, such as motifs and domains. It also describes algorithms for assessing similarity between proteins based on sequence and structure analysis as well as mathematical models, such as hidden Markov models and support vector machines, that are used to represent protein families and classify new instances. The second part covers methods that investigate higher order structure in the protein space through the application of unsupervised learning algorithms, such as clustering and embedding. The book also explores the broader context of proteins. It discusses methods for analyzing gene expression data, predicting protein-protein interactions, elucidating cellular pathways, and reconstructing gene networks. This book provides a coherent and thorough introduction to proteome analysis. It offers rigorous, formal descriptions, along with detailed algorithmic solutions and models. Each chapter includes problem sets from courses taught by the author at Cornell University and the Technion. Software downloads, data sets, and other material are available at biozon.org

This book is concerned with applications of spectroscopic methods to determine protein structure in solution. The methods which are covered include virtually every method which involves the interaction of electromagnetic radiation with proteins in solution and the chapters are written by acknowledged experts from industry and academic institutions. The reader will be able to use this book to decide which spectroscopic techniques will be applicable to the problem he is faced with and be referred to the primary literature which should be most helpful.

New insights into the evolution and nature of proteins

Exploring several distinct approaches, this book describes the methods for comparing protein sequences and protein structures in order to identify homologous relationships and classify proteins and protein domains into evolutionary families. Readers will discover the common features as well as the key philosophical differences underlying the major protein classification systems, including Pfam, Panther, SCOP, and CATH. Moreover, they'll discover how these systems can be used to understand the evolution of protein families as well as understand and predict the degree to which structural and functional information are shared between relatives in a protein family.

Edited and authored by leading international experts, "Protein Families "offers new insights into protein families that are important to medical research as well as protein families that help us understand biological systems and key biological processes such as cell signaling and the immune response. The book is divided into three sections: Section I: Concepts Underlying Protein Family Classification reviews the major strategies for identifying homologous proteins and classifying them into families.Section II: In-Depth Reviews of Protein Families focuses on some fascinating super protein families for which we have substantial amounts of sequence, structural and functional data, making it possible to trace the emergence of functionally diverse relatives.Section III: Review of Protein Families in Important Biological Systems examines protein families associated with a particular biological theme, such as the cytoskeleton.

All chapters are extensively illustrated, including depictions of evolutionary relationships. References at the end of each chapter guide readers to original research papers and reviews in the field.

Covering protein family classification systems alongside detailed descriptions of select protein families, this book offers biochemists, molecular biologists, protein scientists, structural biologists, and bioinformaticians new insight into the evolution and nature of proteins.

Prognostic and Therapeutic Applications of RKIP in Cancer provides updated reviews on the chemistry, signaling, pre-clinical and clinical activities, and role of RKIP expression levels for diagnostics, prognosis and potential interventions. The development of novel compounds and conjugates that selectively induce RKIP expression in cancer open a novel era of new therapeutics and their potential in the treatment of highly resistant cancers and metastases. Edited and written by internationally renowned experts in the field of novel therapeutics for cancer, this book is a valuable source for cancer researchers, medical scientists, clinicians, clinical pharmacologists, and graduate students.

Using a geometric perspective, Protein Geometry, Classification, Topology, and Symmetry reviews and analyzes the structural principals of proteins with the goal of revealing the underlying regularities in their construction. It also reviews computer methods for structure analysis and the automatic comparison and classification of these structures with an analysis of the statistical significance of comparing different shapes. Following an analysis of the current state of protein classification, the authors explore more abstract geometric and topological representations, including the occurrence of knotted topologies. The book concludes with a consideration of the origin of higher-level symmetries in protein structure. The authors focus on simple geometric methods that are deterministic rather than probabilistic and on the more abstract simplifications of protein structure that allow a better understanding of the overall fold of the structure. Most of the methods described in this book have corresponding computer programs. These can be found (as C source code) at the ftp site of the Division of Mathematical Biology at the National Institute for Medical Research. This collection of ideas contains pedagogical material that make it ideal for post-graduate courses as well as new ideas and results essential for researchers investigating protein structures.

Offers coverage of the development of protein purification processes for large-scale commercial operations, and addresses process development, scale-up, applications and mathematical descriptions. Technologies currently used at the commercial scale are covered in depth.

Specialist Periodical Reports provide systematic and detailed review coverage of progress in the major areas of chemical research. Written by experts in their specialist fields the series creates a unique service for the active research chemist, supplying regular critical in-depth accounts of progress in particular areas of chemistry. For over 80 years the Royal Society of Chemistry and its predecessor, the Chemical Society, have been publishing reports charting developments in chemistry, which originally took the form of Annual Reports. However, by 1967 the whole spectrum of chemistry could no longer be contained within one volume and the series Specialist Periodical Reports was born. The Annual Reports themselves still existed but were divided into two, and subsequently three, volumes covering Inorganic, Organic and Physical Chemistry. For more general coverage of the highlights in chemistry they remain a 'must'. Since that time the SPR series has altered according to the fluctuating degree of activity in various fields of chemistry. Some titles have remained unchanged, while others have altered their emphasis along with their titles; some have been combined under a new name whereas others have had to be discontinued. The current list of Specialist Periodical Reports can be seen on the inside flap of this volume.

Lectins have in the past been regarded by many scientists as curious proteins of uncertain structure and specificity that bind to carbohydrates of dubious significance themselves. All this is rapidly changing. The functional importance of glycosylation in cell-cell and cell-pathogen interactions, as well as intracellular events, has been recognized by the explosion of the science of glycobiology. This has been paralleled by the realization that lectins, once they have been well characterized, can be extremely useful tools for exam- ing structural changes in glycosylation and their functional consequences for human pathophysiology. Different lectins vary considerably in their degree of specificity. Some, such as wheatgerm agglutinin, have fairly broad specificity (for glucosamine or sialic acid), whereas others, such as Maackia amurensis, are specific not only for a single carbohydrate, but also for its linkage (2-3 linked sialic acid). Lectins with relatively broad specificity may be very useful as an adjunct to isolation or quantification of soluble glycoproteins, whereas lectins of known, and precise, specificity will be more useful for characterization of carbo- drate structure. We have included an appendix in Lectin Methods and Pro- cols that provides the known specificities of all lectins cited in the text.

"Amino Acid Metabolism, 3rd Edition" covers all aspects of the biochemistry and nutritional biochemistry of the amino acids. Starting with an overview of nitrogen fixation and the incorporation of inorganic nitrogen into amino acids, the book then details other major nitrogenous compounds in micro-organisms, plants and animals. Contents include a discussion of the catabolism of amino acids and other nitrogenous compounds in animals, and the microbiological reactions involved in release of nitrogen gas back into the atmosphere. Mammalian (mainly human) protein and amino acid requirements are considered in detail, and the methods that are used to determine them.

Chapters consider individual amino acids, grouped according to their metabolic origin, and discussing their biosynthesis (in plants and micro-organisms for those that are dietary essentials for human beings), major metabolic roles (mainly in human metabolism) and catabolism (again mainly in human metabolism). There is also discussion of regulatory mechanisms for all these metabolic pathways, and of metabolic and genetic diseases affecting the (human) metabolism of amino acids.

Throughout the book the emphasis is on the nutritional importance of amino acids, integration and control of metabolism and metabolic and other disturbances of relevance to human biochemistry and health. Completely revised edition of this comprehensive text covering all the latest findings in amino acid metabolism researchWritten by an authority in the field Covers new advances in structural biologyClear illustrations of all structures and metabolic pathwaysFull list of recommended further reading for each chapter and bibliography of papers cited in the text

Understanding Biology Using Peptides: Proceedings of the 19th American Peptide Symposium highlights many of the recent developments in peptide science, with a particular emphasis on how these advances are being applied to basic problems in biology and medicine. Specific topics covered include novel synthetic strategies, peptides in biological signaling, post-translational modifications of peptides and proteins, peptide quaternary structure in material science and disease, and peptides as tools in drug discovery.

About the Editor:

Dr. Sylvie Blondelle is an Associate Member in Microbiology/Biochemistry at Mixture Science, Inc., San Diego, CA and Head of the Biosafety Department at the Burnham Institute for Medical Research. Her research focuses on strategies toward the development an HIV vaccine through optimization of peptide immunogens. Dr. Blondelle received her Ph.D. in organic chemistry from the University of Montpellier, France, in 1988.

Driven by the widespread growth of proteomic practices, protein separation techniques have been refined to minimize variability, optimize particular applications, and adapt to user preferences in the analysis of proteins. Separation Methods in Proteomics provides a comprehensive examination of all major separation techniques for proteomics research. Written as a compilation of hands-on methods exemplified by the work of several recognized leaders in the field, this book may serve as a guide for selection of the optimal separation strategies to solve particular biological problems. Recent progress in the development of robust analytical techniques and instrumentation has created the need for good quality biological samples that are subject to analysis. Emphasizing the importance of sample preparation, the book explains how proteomes can be divided into smaller, less complicated "subproteomes" for individual analysis. It also highlights several hybrid approaches that take into account protein interactions. Including applications of the separation methods currently employed in proteomic analyses for both clinical and basic research, Separation Methods in Proteomics contains practical information that can enhance the current and future endeavors of scientists in proteomics, genomics, transcriptomics, biomarker discovery, and drug discovery.