ABPP Methodology: Introduction and Overview, by Matthew B. Nodwell und Stephan A. Sieber Activity-Based Protein Profiling for Natural Product Target Discovery, by Joanna Krysiak und Rolf Breinbauer Photoaffinity Labeling in Activity-Based Protein Profiling, by Paul P. Geurink, Laurette M. Prely, Gijs A. van der Marel, Rainer Bischoff und Herman S. Overkleeft Application of Activity-Based Protein Profiling to the Study of Microbial Pathogenesis, by William P. Heal und Edward W. Tate Functional Analysis of Protein Targets by Metabolomic Approaches, by Yun-Gon Kim und Alan Saghatelian

Recent publications have addressed specific aspects of the endothelins, such as their roles in disease or the importance of endothelin receptors. However, in this book the entire field of endothelins is covered. This includes the pathways of endothelin production and their regulation, the local and systemic actions of endothelins, receptors for the endothelins and the signalling pathways employed, and the involvement of endothelins in a range of diseases. Attention is also paid to the development through chemistry, pharmacology and toxicology of endothelin antagonists and endothelin-converting enzyme inhibitors, with mention of all the important members of these drug classes. This leads to well-rounded discussions of the potential therapeutic benefit of endothelin inhibitors.

The ubiquitin-proteasome system (UPS) and ubiquitin-related modifiers are not only involved in cellular protein quality control but also in the regulation of many fundamental cellular processes/pathways as well as in their disease-relevant aberrations. Ubiquitin Family Modifiers and Proteasome: Reviews and Protocols presents both novel developments in UPS research and important methods related to the main recent advances in the field of ubiquitin family modifiers. Divided into five convenient sections, this volume focuses on the enzymology and substrate identification of ubiquitin family modifiers, the recognition and chain formation of these modifiers, the analysis of proteasome biogenesis and function, protein quality control, and finally the use of small molecules and strategies to study or manipulate the function of the UPS and of ubiquitin family modifiers, respectively. 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 protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Ubiquitin Family Modifiers and Proteasome: Reviews and Protocols will be of great use to investigators and students engaged in both basic and applied research in life sciences.

Bringing together a wide variety of examples of functional amyloid in a single volume, this book explores the importance of amyloid fibrils in fungi, bacteria, algae, invertebrate, and vertebrate animals for providing environmental protection, structural integrity, and regulating biochemical processes. It highlights many of the extraordinary examples of functional amyloid found to date. It, therefore, provides an exciting perspective for the study of amyloid deposits as important and useful protein structures widespread in nature.

Knowledge about protein tertiary structure can guide experiments, assist in the understanding of structure-function relationships, and aid the design of new therapeutics for disease. Homology modeling is an in silico method that predicts the tertiary structure of an amino acid sequence based on a homologous experimentally determined structure. In, Homology Modelling: Methods and Protocols experts in the field describe each homology modeling step from first principles, provide case studies for challenging modeling targets and describe methods for the prediction of how other molecules such as drugs can interact with the protein. 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, Homology Modelling: Methods and Protocols guides scientists in the available homology modeling methods.

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.

Sequencing projects have revealed the presence of at least several hundred receptor kinases in a typical plant genome. Receptor kinases are therefore the largest family of primary signal transducers in plants, and their abundance suggests an immense signaling network that we have only just begun to uncover. Recent research findings indicate that individual receptor kinases fulfill important roles in growth and development, in the recognition of pathogens and symbionts or, in a few examples, in both growth and defense. This volume will focus on the roles of receptor kinases, their signaling pathways, and the ways in which these important signaling proteins are regulated.

Now recognized as a reservoir for growth factors and cytokines modulating cell activation status and turnover, proteoglycans have stimulated great amount of interest and research. In Proteoglycans: Methods and Protocols, experts in the field examine issues of basic concepts and up-to-date analysis methods for proteoglycan and glycosaminoglycan (GAG) at the protein and saccharide levels. The volume continues with the multifunctional aspect of proteoglycans, highlighted through three relevant examples of proteoglycans with highly different strucutures: serglycin, aggrecan, and heparin sulphate proteoglycans. The final chapter describes proteoglycan involvement in the pathogenesis of various disorders and their potential therapeutic value in osteo-articular diseases. 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 tips on troubleshooting and avoiding known pitfalls. Authoritative and accessible, Proteoglycans: Methods and Protocols is an ideal guide for scientists attempting to pursue further research in this vital and complex area of study.

Amyloid-forming proteins are implicated in over 30 human diseases. The proteins involved in each disease have unrelated sequences and dissimilar native structures, but they all undergo conformational alterations to form fibrillar polymers. The fibrillar assemblies accumulate progressively into disease-specific lesions in vivo. Substantial evidence suggests these lesions are the end state of aberrant protein folding whereas the actual disease-causing culprits likely are soluble, non-fibrillar assemblies preceding the aggregates. The non-fibrillar protein assemblies range from small, low-order oligomers to spherical, annular, and protofibrillar species. Oligomeric species are believed to mediate various pathogenic mechanisms that lead to cellular dysfunction, cytotoxicity, and cell loss, eventuating in disease-specific degeneration and systemic morbidity. The particular pathologies thus are determined by the afflicted cell types, organs, systems, and the proteins involved. Evidence suggests that the oligomeric species may share structural features and possibly common mechanisms of action. In many cases, the structure function interrelationships amongst the various protein assemblies described in vitro are still elusive. Deciphering these intricate structure function correlations will help understanding a complex array of pathogenic mechanisms, some of which may be common across different diseases albeit affecting different cell types and systems."

Fluorescent proteins are intimately connected to research in the life sciences. Tagging of gene products with fluorescent proteins has revolutionized all areas of biosciences, ranging from fundamental biochemistry to clinical oncology, to environmental research. The discovery of the Green Fluorescent Protein, its first, seminal application and the ingenious development of a broad palette of fluorescence proteins of other colours, was consequently recognised with the Nobel Prize for Chemistry in 2008.

"Fluorescent Proteins II" highlights the physicochemical and biophysical aspects of fluorescent protein technology beyond imaging. It is tailored to meet the needs of physicists, chemists and biologists who are interested in the fundamental properties of fluorescent proteins, while also focussing on specific applications. The implementations described are cutting-edge studies and exemplify how the physical and chemical properties of fluorescent proteins can stimulate novel findings in life sciences.

Protein-protein interactions (PPIs) are strongly predictive of functional relationships among proteins in virtually all processes that take place in the living cell. Therefore, the comprehensive exploration of interactome networks is one of the major goals in systems biology. The aim of Two Hybrid Technologies: Methods and Protocols is to provide a compendium of state-of-the art protocols for the investigation of binary PPIs with the classical yeast two-hybrid (Y2H) approach, Y2H variants and other in vivo methods for PPI mapping. Divided into two convenient sections, the first gives a survey of protocols that are currently employed for Y2H high-throughput screens by different expert labs in the field. Rather than detailing the principles of screening, which have been described previously, the focus is on different implementations of Y2H interactome mapping. The second section of the book considers innovative PPI detection methods that have the potential to emerge as alternative high-throughput methodologies. 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, Two Hybrid Technologies: Methods and Protocols supplies researchers with a comprehensive toolbox for the identification of biologically relevant protein interactions.

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.

This volume is a collection of the contributions presented at the 42nd Erice Crystallographic Course whose main objective was to train the younger generation on advanced methods and techniques for examining structural and dynamic aspects of biological macromolecules. The papers review the techniques used to study protein assemblies and their dynamics, including X-ray diffraction and scattering, electron cryo-electron microscopy, electro nanospray mass spectrometry, NMR, protein docking and molecular dynamics. A key theme throughout the book is the dependence of modern structural science on multiple experimental and computational techniques, and it is the development of these techniques and their integration that will take us forward in the future.

This volume is a collection of the contributions presented at the 42nd Erice Crystallographic Course whose main objective was to train the younger generation on advanced methods and techniques for examining structural and dynamic aspects of biological macromolecules. The papers review the techniques used to study protein assemblies and their dynamics, including X-ray diffraction and scattering, electron cryo-electron microscopy, electro nanospray mass spectrometry, NMR, protein docking and molecular dynamics. A key theme throughout the book is the dependence of modern structural science on multiple experimental and computational techniques, and it is the development of these techniques and their integration that will take us forward in the future.

Amino Acid Analysis (AAA) is an integral part of analytical biochemistry. In a relatively short time, the variety of AAA methods has evolved dramatically with more methods shifting to the use of mass spectrometry (MS) as a detection method. Another new aspect is miniaturization. However, most importantly, AAA in this day and age should be viewed in the context of Metabolomics as a part of Systems Biology. Amino Acid Analysis: Methods and Protocols presents a broad spectrum of all available methods allowing for readers to choose the method that most suits their particular laboratory set-up and analytical needs. In this volume, a reader can find chapters describing general as well as specific approaches to the sample preparation. A number of chapters describe specific applications of AAA in clinical chemistry as well as in food analysis, microbiology, marine biology, drug metabolism, even archeology. Separate chapters are devoted to the application of AAA for protein quantitation and chiral AAA. 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, Amino Acid Analysis: Methods and Protocols provides crucial techniques that can be applied across multiple disciplines by anyone involved in biomedical research or life sciences.

Haemocyanin was first recognised as a respiratory pigment by P. Bert in 1867. Over the years the haemocyanins have attracted attention as macromolecules and copper-contain- ing respiratory proteins. The early functional studies of A.C. Redfield (Biol. Rev. 9, 176, 1934) and the ultracen- trifuge work of I.B. Ericksson-Quensel and T. Svedberg (Biol. Bull. 21, 498, 1936) come easily to mind. In recent years haemocyanin studies have come to the forefront with the work of the Ghirettis (Padova), R. Lontie (Louven), E.F.J. Van Bruggen (Groningen) and Joe and Celia Bonaven- tura (Beaufort) to mention but a few of the number of able investigators coming to this field from diverse disciplines. It is hoped that this book presents a fair cross section of current workers and work on haemocyanin. It is the sec- ond collection of haemocyanin studies to be published after a Workshop on the Structure and Function of Haemocyanin. The first haemocyanin meeting was held by Professors F. "Ghiro" Ghiretti and Anna Ghiretti-Magaldi in Naples in 1966. Subsequent meetings have been held at Groningen (1970), Louven (1971) and Padova (1974). Current interest in haemocyanin can be judged by the scope and breadth of the papers from the Malta meeting and pre- sented here. In organising the Malta meeting I would like to thank Dr. John Tooze of the European Molecular Biology Organisation and my friend and colleague, Professor Mauri- zio Brunori for their continuous help and support.

SELDI is distinct from other TOF-MS technologies in that it couples features of chromatography and mass spectrometry, facilitating analyte enrichment and sample cleanup on an array surface. In the growing field of proteomics, SELDI technology has been widely used for biomarker discovery and characterization in diverse applications including diagnostics, drug development, and basic research. SELDI-based biomarker studies can typically be divided into four phases: discovery, validation, purification and identification, and assay development. SELDI-TOF Mass Spectrometry: Methods and Protocols provides an overview of the current applications of SELDI-TOF MS (surface enhanced laser desorption/ionization time-of-flight mass spectrometry), with an emphasis on study and experimental design, data analysis and interpretation, and assay development. 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, SELDI-TOF Mass Spectrometry: Methods and Protocols will provide information on optimizing study design, experimental protocols, and data analysis and interpretation to yield robust biomarkers and biomarker assays, using examples from different disease areas.

Display technologies have become a very powerful way of generating therapeutic lead molecules and specific reagents for increasing our understanding of biology; however, despite being first described shortly after phage display, the use of ribosome display and related methods have been much less widespread. Since this is in part due to the complexity of the methods, "Ribosome Display and Related Technologies: Methods and Protocols" seeks to extend their use by collecting expert contributions describing these detailed protocols. The protocols described range from well-established methods that have been used for a decade to generate high affinity antibodies, which are already in the clinic, to methods that are in their early stages of application such as display of peptides incorporating non-canonical amino acids. 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.

Invaluable and easy to use, "Ribosome Display and Related Technologies: Methods and Protocols" will be of great benefit to those with general molecular biology or protein engineering experience who wish to select peptides or proteins by display, those with phage display experience who would benefit from the application of ribosome display, as well as those with some ribosome display experience who would like to expand the range of applications to which they are applying the technology."

The multidisciplinary science of chemical proteomics studies how small molecules of synthetic or natural origin bind to proteins and modulate their function. In Chemical Proteomics: Methods and Protocols, expert researchers in the field provide key techniques to investigate chemical proteomics focusing on analytical strategies, how probes are generated, techniques for the discovery of small molecule targets and the probing of target function, and small molecule ligand and drug discovery. 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, Chemical Proteomics : Methods and Protocols seeks to provide methodologies that will contribute to a wider application of chemical proteomics methods in biochemical and cell biological laboratories.

Despite considerable variability within the scientific community, allosteric regulation can best be defined functionally as how a macromolecule binds one ligand differently when a second ligand is or is not pre-bound to the macromolecule, which constitutes a vital aspect of protein structure/function. In "Allostery: Methods and Protocols," expert researchers in the field provide key techniques to investigate this biological phenomenon. Focusing on heterotropic systems with some coverage of homotropic systems, this volume covers the monitoring of allosteric function, allosteric conformational changes, and allosteric changes in protein dynamics/sub-population distribution, as well as topics such as macromolecular and ligand engineering of allosteric functions and computational aids in the study of allostery. Written in the highly successful "Methods in Molecular Biology" 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, "Allostery: Methods and Protocols" aids scientists in continuing to study ligand-induced, through-protein effects on protein function (ligand binding/catalysis), a phenomenon that is well recognized through the history of the life sciences and very poorly understood at the molecular level."

Fluorescent proteins are intimately connected to research in the life sciences. Tagging of gene products with fluorescent proteins has revolutionized all areas of biosciences, ranging from fundamental biochemistry to clinical oncology, to environmental research. The discovery of the Green Fluorescent Protein, its first, seminal application and the ingenious development of a broad palette of fluorescence proteins of other colours, was consequently recognised with the Nobel Prize for Chemistry in 2008.

"Fluorescent Proteins I" is devoted to the basic photophysical and photochemical aspects of fluorescent protein technology. Experienced experts highlight colour tuning, the exploration of switching phenomena and respective methods for their investigation. The book provides a thorough understanding of primary molecular processes allowing the design of fluorescent proteins for specific applications.

The present volume contains the proceedings of the 23rd Mos bach Colloquium on "Protein-Protein Interactions." It includes the paper by Prof. S. LIFSON who unfortunately was unable to attend. Discussions were included whenever possible in order to make the complete proceedings accessible to the future reader. The colloquium was generously supported by the firms who are corporate members of the Gesellschaft fur Biologische Chemie. Dr. W. B. GRATZER and Dr. L. JAENICKE, as weIl as some of the invited speakers, were a tremendous help in preparing the program and we wish to thank them for their helpfuI suggestions and advice. Our appreciation extends to Prof. E. AUHAGEN and Prof. H. GIBIAN, and their secretaries, who shared the burden of organizing the colloquium. In addition, we wish to express our gratitude to Springer-Ver Iag, and especially to Dr. H. MAYER-KAUPP for editoriaI help and for the rapid publication of this volume. We hope that the reader, Iike the audience in the Mosbach market hall, will gain an impression of the present state of protein research and of the way in which intermolecular interactions hold specific implications for the structure and function of proteins. Regensburg and Wurzburg, autumn 1972 R. JAENICKE E. HELMREICH Contents Introduction. R. JAENICKE (Regensburg) 1 Molecular Forees. S. LIFSON (Rehovot) ................... 3 Structure, Function and Dynamics of a ReguIatory Enzyme: Aspartate Transcarbamylase. H. K. SCHACHMAN (Berkeley) 17 Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54 . . . . . . . . . . . . . ."

Plasma lipoproteins constitute a unique macromolecular system of lipid-protein complexes responsible for the transport of lipids from their sites of origin to their sites of utilization either as metabolic fuel or as structural components of cell membranes. Although studies on the role of lipoproteins in the mechanism of lipid transport are meritorious in their own right, the ever-increasing interest in chemical and functional properties of this remarkable class of conjugated proteins stems from the impressive evidence of their direct involvement in the genesis and develop ment of atherosclerotic lesions. The initial emphasis on neutral lipids and phospholipids as the most characteristic constituents of operationally defined lipoprotein classes has shifted in recent years to their protein moieties or apolipoproteins. The discovery of a number of apolipoproteins and characterization of familial hypolipoproteinemias as apolipoprotein deficiency disorders indicated that apolipoproteins play an essential role in maintaining the structural stability and integrity of lipoprotein particles. In addition to their role in the formation of lipoproteins, apolipoproteins were shown to perform a variety of functions in metabolic conversion of lipoproteins and their interactions with cellular surfaces. Results from several laboratories have demonstrated that the chemical and metabolic heterogeneity of operationally-defined lipoprotein classes is due to the presence of several discrete lipoprotein particles with similar physical properties but different and characteristic apolipoprotein composition. Thus, the apolipoproteins have emerged not only as essential structural and functional constituents of lipoproteins but also as unique chemical markers for identifying and classifying lipoprotein particles."

We are proud to present Volume 3 of Biological Magnetic Resonance, a series that has met with praise from the scientific community. This volume covers the new applications of various multiple irradia- tion techniques to the NMR of biomolecules; the chapter of Keller and Wuthrich describes much of the technique and its applications to hemo- proteins. The ESR of some hemoproteins in the single crystal is described by Chien and Dickinson, who also include discussions of techniques and methods for single-crystal ESR of paramagnetically intrinsic and spin- labeled protein crystals. Mims and Peisach describe the latest applications and results in electron spin echo spectroscopy of several metalloproteins. Two ESR spin probe techniques are reviewed. Chasteen describes the methods and applications of vanadyl(JV) to several systems. Ohnishi and Tokutomi describe studies of phase separations in mixed and model mem- branes by the nitroxide spin probe technique. We have been successful in continuing to provide topics that are timely and experimentally informative with a heavy emphasis on biolo- gically relevant applications. We thank our colleagues in the scientific com- munity for their suggestions on future coverage-we will remain receptive to future suggestions and comments on this series. A tentative topic list for forthcoming volumes is given on the following pages.