This second edition presents an up-to-date chapters describing the most relevant and novel techniques employed to study the opioid receptors. Chapters detail transcriptional and post-transcriptional analysis, cellular detection of opioid receptors, analysis of signaling events modulated by opioid receptors, model systems to studying opioid receptor-mediated functions, and behavioral effects mediated by opioid receptors. 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, Opioid Receptors: Methods and Protocols, Second Edition aims to ensure successful results in the further study of this vital field.

Principles of Proteomics, Second Edition, provides a concise and user-friendly introduction to the diverse technologies used for the large-scale analysis of proteins, as well as their applications, and their impact in areas such as drug discovery, agriculture, and the fight against disease. Proteomics is a fast-advancing field in which researchers seek to capture all the proteins in the cell and characterize them in ever more detail. Principles of Proteomics has been fully updated to reflect the most recent developments in the field without losing its focus on the underlying principles. With worked examples, case studies profiling both established and emerging technologies, and further reading lists for each chapter, Principles of Proteomics is an ideal introduction for students, researchers and those working in the industry.

One of the phylogenetically oldest hormones, somatostatin is a regulatory peptide with remarkable characteristics. It is a nonclassical neurotransmitter discovered less than 20 years ago both in the central nervous system and in the gastroenteropancreatic system. It regulates the secretion of both pituitary and digestive hormones, it ensures nutrient homeostasis and it has therapeutic uses. This volume deals withall these aspects.

0ne 0fthe many 1mpact5 0frec0m61nant DNA techn0109y 0ver the 1a5t 15 year5 ha5 6een a 5tr0n91y refre5hed 1ntere5t 1n meth0d5 f0r the 5eparat10n and pur1f1cat10n 0f pr0te1n5.7h15 1ntere5t ha5 enc- pa55ed n0t 0n1y ana1yt1ca1 5eparat10n5, 6ut a150 5ma11- and 1ar9e-5ca1e preparat1ve meth0d5 d1rected t0 60th pure and app11ed re5earch thr0u9h0ut 610109y and 610med1c1ne. Many 0f the new 0r 5u65tant1a11y m0d1f1ed techn14ue5 deve1- 0ped have 6een rep0rted 1n the 11terature, 6ut a 5uff1c1ency 0f deta11ed pract1ca1 he1p 1n e5ta6115h1n9 the5e meth0d5 f0r the f1r5t t1me 1n a new 1a60rat0ry ha5 0ften 6een d1ff1cu1t t0 f1nd. W1th the5e pr061em5 1n m1nd, we expect that Pract1ca1Pr0te1n Chr0mat09raphy, de519ned a5 a key v01ume 1n the Meth0d5 1n M01ecu1ar 810109y 5er1e5, w111 pr0v1de c0nc15e pract1ca1 he1p t0 th05e carry1n9 0ut new techn14ue5 f0r the f1r5t t1me. Each chapter ha5 6een wr1tten 6y expert auth0r5 kn0wn t0 have d1rect and re9u1ar pract1ca1 exper1ence w1th the1r ch05en techn14ue5. 7he 5tructure 0feach chapter 15 de519ned t0 make 1t ea5y f0r a w0rker new t0 the meth0d t0 f0110w 1t t0 an effect1ve c0nc1u510n. An 1nt- duct10n treat5 the the0ry 6eh1nd the meth0d 6e1n9 de5cr16ed. 7he Mater1a15 and Meth0d5 5ect10n5 a110w the reader t0 prepare f0r, and then perf0rm techn14ue5 1n a rat10na1 5tepw15e manner. 7he N0te5 5ect10n5 pr0v1de the 50rt 0f6ack9r0und h1nt5 and tr1ck5 that are 50 0ften e55ent1a1 f0r 5ucce55, 6ut are rare1y rep0rted 1n the 11terature.

A Safety Considerations Many techniques described here involve a number of hazards, such as high electrical current and voltage, radioactivity and highly toxic chemicals. It is absolutely essential that the instructions of equipment manufacturers be followed, and that particular attention be paid to the local and federal safety regulations. B Introduction The expression of prokaryotic and eukaryotic genes has been shown most often to be regulated at the level of mRNA synthesis. Thanks to the rapid development of methods for dissecting DNA sequences, cis-acting regulatory elements such as promoters and enhancers have been recognised. More recently, the widely expressed intuition that discrete sequences within these elements constitute binding sites for sequence-specific binding proteins has been confirmed, especially through the use of "footprinting" assays (for examples, Galas and Schmitz, 1978). This and similar assays have already resulted in the recognition, isolation and analysis of DNA-bind ing proteins for several genes. Excellent reviews exist of the structural studies on these transcription regulatory proteins and related DNA elements (for example, Glover, 1989 and Johnson and McKnight, 1989), to which the reader is referred for detailed information. To set the scene for applications of the techniques described in this volume, only the barest outline of previous studies is presented here. Protein-DNA interactions are dependent on very specific tertiary configurations of the binding protein which allow the closest contact with the DNA helix.

There is increasing evidence for the clinicial value of the apo lipoprotein measurements. Besides cholesterol in plasma and li poprotein fractions, which is currently used as an indicator of cardiovascular risk, the measurement of the AI and B apolipopro teins can provide additional information about the patients' clinical status. Several studies show that apo B is higher and apo AI is lower in patients with angiographically documented coronary heart dis ease than in symptomatic patients without coronary heart disease. Moreover, discriminant analysis indicated that the concentration of Apo AI and B in plasma are better discriminators than lipo protein cholesterol for identifying patients with coronary heart disease. In some studies the apo Bjapo AI ratio appears to be a more powerful predictor than individual lipoproteins. In a recent study carried out in men, apolipoproteins AI and B were better correlated with the severity of cardiovascular disease than HDL and LDL cholesterol. The predictive power of apolipoproteins could however not be demonstrated in all studies and the value of apolipoprotein measurements in the field of clinical chemistry is still controversial. This is probably due to discrepancies between the results of various studies, arising from differences in the type of immunoassays, the lack of universal reference materials, differ ences between study protocols, variations in the selection of patients and in the grading and interpretation of coronary lesions."

It is our great pleasure to introduce the Proceedings of the Twenty-First European Peptide Symposium, held in Platja d' Aro, Spain, September 2-8, 1990. Over seven hundred scientists from nearly thirty countries, mostly European but also from the Americas, Australia and Japan, assembled at the largest-to-date in a series of European Peptide Symposia to present and discuss their recent findings in the field of peptides. Whenever scientists meet, a good deal of their interaction cannot be appropri ately recorded in book form. Fruitful early morning dialogues go regrettably unrecorded or, at most, precariously scribbled on ephemeral breakfast napkins. Many a brilliant late night debate is bound to fade away through the mists of a noisy discotheque. Alas, this book will unfortunately ignore these potentially splendid contributions to science! No effort has been made either to register the discussions that followed the oral presentations of the Symposium, or the two open sessions on special topics of general interest. Spontaneity, the main appeal of oral discussion, is somewhat lost, we believe, on literal transcription. Once all these non-recordable forms of scientific communication are discounted one is left with the core of the Symposium, i.e., its scientific communications, both in oral and poster form, and it is to these that the book you have in your hands is devoted.

This book reviews the principles of design and examples of successful implementation of proteinkinase inhibitors (PKI), and offers a comprehensive and authoritative overview of the history and latest developments in the field. Chapters written by experts from industry and academia cover the function, structure and topology of Proteinkinases, molecular modelling, disclose how to achieve high level of selectivity for kinase inhibitors, and exploit kinase inhibitors for cancer treatment. Particular attention is given to Inhibitors of c-Jun N-terminal kinase 3, and to covalent Janus Kinase 3 Inhibitors. A case study on Receptor Tyrosine Kinases EGFR, VEGFR, PDGFR is also presented in this book. Given its breath, this book will appeal to medicinal chemists, students, researchers and professionals alike.

Techniques in the neurosciences are evolving rapidly. There are currently very few volumes dedicated to the methodology - ployed by neuroscrentists, and those that are available often seem either out of date or limited in scope. This series is about the methods most widely used by modern-day neuroscientists and is written by their colleagues who are practicing experts. Volume 1 will be useful to all neuroscientists since rt concerns those procedures used routmely across the wrdest range of s- disciplines. Collecting these general techniques together in a single volume strikes us not only as a service, but will no doubt prove of exceptronal utrlitarran value as well. Volumes 2 and 3 describe current procedures for the analyses of amines and their metabolites and of amino acids, respectively. These collections will clearly be of value to all neuroscrentists working m or contemplating research m these fields. Similar reasons exist for Volume 4 on receptor binding techniques, since experimental details are provided for many types of hgand-receptor binding, including chapters on general prin- ples, drug discovery and development, and a most useful app- dix on computer programs for Scatchard, nonlinear, and compe- tive displacement analyses. Volume 5 provides procedures for the assessment of enzymes involved in neurotransmitter synthesis and catabolism. Volumes in the NEUROMETHODS series will be useful to neurochemists, -pharmacologists, -physiologrsts, -anatomrsts, psychopharmacologists, psychiatrists, neurologists, and chemists (organic, analytical, pharmaceutical, medicinal); in fact, everyone involved m the neuroscrences, both basic and clmical.

Methods in Protein Sequence Analysis -1986 brings together reports of the most recent methodology available to protein chemists for studying the molecular detail of proteins. The papers in this volume constitute the proceedings of the Sixth International Conference on Methods in Protein Sequence Analysis, which was held at the University of Washington in Seattle, Washington on August 17-21, 1986. This series of conferences has taken place during a period when new techniques in protein chemistry and molecular biology have enabled not only exploration of the control of protein function, but also deduction of the genetic origin of proteins, and labo ratory generation of rare protein molecules for therapeu tic and commercial use. The current reports are focused on the means by which experimental questions can be answered rather than on the biological implications in specific systems. The scope of the meeting was quite broad, empha sizing microanalytical techniques and the relative merits of DNA sequencing, mass spectrometry and more tradi tional degradation techniques. A highlight of the meeting was the Qrowing awareness of the role of mass spec trometry In the analysis of proteins. The complementarity of protein sequencing and DNA sequencing techniques was apparent throughout the discussions and several papers dealt with the strategy of obtaining sequence in formation from small amounts of protein in order that ap propriate oligonucleotide probes could be constructed and the encoding nucleic acids se. quenced and manipu lated."

Milestones in the techniques and methodology of polypeptide structure determination include the determination of the sequence of insulin by Sanger in 1951 (I) and the introduction of the repeti tive degradation of proteins with phenylisothiocyanate by Edman in 1959 (2). The automation of Edman chemistry (3) played a major role in the determination of polypeptide structures. Important modifications of Edman chemistry include the solid-phase approach by Laursen in 1971 (4) and the use of modified Edman reagents such as 4-N, N-dimethylaminoazobenzene-4'-isothiocy- ate (DABITC) for manual sequencing by Chang et al. (5) in 1976. A second major breakthrough in the analysis of polypeptides was automated amino acid analysis described by Spackman et al. in 1958 (6). However, during the period from 1975 to 1980, it became increasingly clear that the amount of material required for struc tural analysis was more than could be easily isolated for the vast majority of proteins. The field was criticized for its lack of sensitive techniques for the analysis of growth factors, immune modulators, membrane receptors, and peptide hormones. In addition, very little had been done to modernize and improve the original instruments introduced in the mid-1960s. The first indications of improved instrumentation for Edman chemistry came from Wittmann-Liebold's laboratory (7), followed by the introduction of a "micro" sequencer by Hunkapiller and Hood in 1978 (8). The movement toward improved instrumentation culminated in the "gas"-phase sequencer of Hewick et al. (9) in 1981."

Techniques in the neurosciences are evolving rapidly. There are currently very few volumes dedicated to the methodology - ployed by neurosclentlsts, and those that are available often seem either out of date or limited m scope. This series is about the methods most widely used by modern-day neurosclentists and 1s wrltten by their colleagues who are practicing experts. Volume 1 will be useful to all neurosclentists since It concerns those procedures used routinely across the widest range of subdisciplines. Collecting these general techniques together m a single volume strikes us not only as a service, but will no doubt prove of exceptIona utilitarian value as well. Volumes 2 and 3 - scribe all current procedures for the analyses of amines and their metabolltes and of ammo aads, respectively. These collections will clearly be of value to all neuroscientlsts working m or contemplating research m these fields. Similar reasons exist for Volume 4 on receptor binding techniques since experimental - tails are provided for all types of llgand-receptor bmdmg, including chapters on general prmciples, drug discovery and - velopment, and a most useful appendix on computer programs for Scatchard, nonlinear, and competitive displacement analyses. Volume 5 provides procedures for the assessment of enzymes - volved m biogenic amme synthesis and catabolism. Volumes in the NELJROMETHODS series will be useful to neurochemists, -pharmacologists, -physiologists, -anatomlsts, psychopharmacologists, psychlatnsts, neurologists, and chemists (organic, analytical, pharmaceutical, medicinal), in fact, everyone involved m the neurosclences, both basic and clinical.

In recent years there has been a tremendous increase in our understanding of the functioning of the cell at the molecular level. This has been achieved in the main by the invention and development of new methodology, parti- larly in that area generally referred to as "'genetic en- neering." While this revolution has been taking place in the field of nucleic acids research, the protein chemist has at the same time developed fresh methodology to keep pace with the requirements of present day molecular bi- ogy. Today's molecular biologist can no longer be content with being an expert in one particular area alone. He/she needs to be equally competent in the laboratory at h- dling DNA, RNA, and proteins, moving from one area to another as required by the problem he/she is trying to solve. Although many of the new techniques in molecular biology are relatively easy to master, it is often difficult for a researcher to obtain all the relevant information nec- sary for setting up and successfully applying a new te- nique. Information is of course available in the research l- erature, but this often lacks the depth of description that the new user requires. This requirement for in-depth pr- tical details has become apparent by the considerable - mand for places on our Molecular Biology Workshops held at Hatfield each summer.

In recent years there has been a tremendous increase in our understanding of the functioning of the cell at the molecular level. This has been achieved in the main by the invention and development of new methodology, parti- larly in that area generally referred to as "'genetic en- neering." While this revolution has been taking place in the field of nucleic acids research, the protein chemist has at the same time developed fresh methodology to keep pace with the requirements of present day molecular bi- ogy. Today's molecular biologist can no longer be content with being an expert in one particular area alone. He/she needs to be equally competent in the laboratory at h- dling DNA, RNA, and proteins, moving from one area to another as required by the problem he/she is trying to solve. Although many of the new techniques in molecular biology are relatively easy to master, it is often difficult for a researcher to obtain all the relevant information nec- sary for setting up and successfully applying a new te- nique. Information is of course available in the research l- erature, but this often lacks the depth of description that the new user requires. This requirement for in-depth pr- tical details has become apparent by the considerable - mand for places on our Molecular Biology Workshops held at Hatfield each summer.

During the past decade we have witnessed several major dis coveries in the area of protein synthesis and post-translational modification of protein molecules. In this volume, many of the lat est research developments in these fields are reported by the dis tinguished international group of scientists who presented their state-of-the-art results at the 13th Linderstr0m-Lang Conference held at God0Ysund, Norway, June 14-18, 1983. We feel that the presentation here of so wide a variety of articles on both the molecular and the cellular aspects of protein synthesis will be of considerable value to many scientists working in the area who were unable to attend, as well as to many who are active in related areas. In addition to the research papers, the contents of the six scientific sessions held during the conference have been summarized by the respective session chairmen. These individual summaries provide insightful syntheses of all the recent progress in each field, identify which current problems remain of special inter est, and suggest what the future may hold in the several areas of protein synthesis research covered. Though this volume obviously cannot provide a complete survey of all important ongoing research on the molecular and cellular biology of translational and post-translational events, we are confi dent that it will facilitate a much better understanding of many im portant contemporary problems in research on protein synthesis, including cell differentiation, translational accuracy, protein modifi cation, intracellular transport, and membrane turnover."

Methods in Protein Sequence Analysis contains an intensely prac tical account of all the new methodology available to scientists carrying out protein and peptide sequencing studies. Many of the striking advances in fields as diverse as immunology, cell motility, and neurochemistry have in fact been fueled by our ever more powerful ability to determine the sequences and structures of key proteins and peptides. It is our hope that the rich array of tech niques and methods for sequencing proteins discussed in this volume-methods that generate much of the information crucial to progress in modern biology-will now become accessible to all who can benefit from them. The papers of the present volume constitute the Proceedings of the IVth International Conference on Methods in Protein Se quence Analysis, which was held at Brookhaven National Labo ratory, Upton, NY, September 21-25, 1981. It was the most recent in a series of biennial conferences, the previous one having been held in Heidelberg, GFR, in 1979. The series was originated by Richard Laursen, and initially dealt with one aspect of the field, solid-phase sequencing. The scope of the meeting was very broad and among the many aspects of protein sequencing discussed were: instrumentation, strategy, chemicals, mass spectrometry, cleavage of proteins and separation of peptides, and solid, liquid, manual, and even "gas phase" sequencing."

The handling and analysis of data generated by proteomics investigations represent a challenge for computer scientists, biostatisticians, and biologists to develop tools for storing, retrieving, visualizing, and analyzing genomic data. Informatics in Proteomics examines the ongoing advances in the application of bioinformatics to proteomics research and analysis. Through computer simulations, scientists can determine more about how diseases affect cells, predict how various drug interventions would work, and ultimately use proteins as therapeutic targets. This book first addresses the infrastructure needed for public protein databases. It discusses information management systems and user interfaces for storage, retrieval, and visualization of the data as well as issues surrounding data standardization and integration of protein sequences recorded in the last two decades. The authors subsequently examine the application of statistical and bioinformatic tools to data analysis, data presentation, and data mining. They discuss the implementation of algorithms, statistical methods, and computer applications that facilitate pattern recognition and biomarker discovery by integrating data from multiple sources. This book offers a well-rounded resource of informatic approaches to data storage, retrieval, and protein analysis as well as application-specific bioinformatic tools that can be used in disease detection, diagnosis, and treatment. Informatics in Proteomics captures the current state-of-the-art and provides a valuable foundation for future directions.

This text offers in-depth perspectives on every aspect of protein structure identification, assessment, characterization, and utilization, for a clear understanding of the diversity of protein shapes, variations in protein function, and structure-based drug design. The authors cover numerous high-throughput technologies as well as computational methods to study protein structures and residues. A valuable reference, this book reflects current trends in the effort to solve new structures arising from genome initiatives, details methods to detect and identify errors in the prediction of protein structural models, and outlines challenges in the conversion of routine processes into high-throughput platforms.

While the genomic revolution has quickly led to the deposit of more than 30,000 structures in the protein data bank (PDB), less than one percent of those contributions represent membrane proteins despite the fact that membrane proteins constitute some 20 percent of all proteins. This discrepancy becomes significantly troublesome when it is coupled with the fact that 60 percent of current drugs are based on targeting this group of proteins, a trend that does not seem likely to reverse. Structural Genomics on Membrane Proteins provides an excellent overview on novel research in bioinformatics and modeling on membranes, as well as the latest technological developments being employed in expression, purification, and crystallography to obtain high-resolution structures on membrane proteins. This cutting-edge work also explains the difficulties facing researchers-both technical and ethical-that have slowed the process. Structural Genomics on Membrane Proteins provides researchers with an unprecedented look at the novel technologies that will ultimately allow them to conquer the last frontier in structural biology, leading to accelerated breakthroughs in drug discovery.

The elucidation of the ultrastructure, cytochemistry and reactivity of cutaneous recep- tors is of particular importance in the unsolved biological problem of sensory transduc- tion. Topographically and with regard to evolution, cutaneous receptors are the most widespread primary sense organs. Because of their simple structure they are particular- ly useful in the study of the functional morphology of receptor end-organs. Research into receptor structures in classic neuromorphology revealed magnificent patterns and pictures of receptors, showing their great diversity and their presence in all human and animal systems. Since the end of the sixties, ultrastructural studies have led to the discovery of a large amount of new information. The findings concerning the cellular organization of the inner core (pease and Quilliam, 1957), axon terminals (Cauna and Ross, 1960) and capsule cells (Polacek and Mazanek, 1966) of encapsulated receptors have opened up new possibilities for a revaluation of receptor cell units and their relationships.

Meeting the need for a book on developing and using new methods to investigate membrane proteins, this is the first of its kind to present the full range of novel techniques in one resource. Top researchers from around the world focus on the physical principles exploited in the different techniques, and provide examples of how these can bring about important new insights.

Following an introduction, further sections discuss structural approaches, molecular interaction and large assemblies, dynamics and spectroscopies, finishing off with an exploration of structure-function relationships in whole cells.

This highly readable textbook serves as a concise and engaging primer to the emerging field of antibody engineering and its various applications. It introduces readers to the basic science and molecular structure of antibodies, and explores how to characterize and engineer them. Readers will find an overview of the latest methods in antibody identification, improvement and biochemical engineering. Furthermore, alternative antibody formats and bispecific antibodies are discussed. The book's content is based on lectures for the specializations "Protein Engineering" and "Medical Biotechnology" within the Master's curriculum in "Biotechnology." The lectures have been held at the University of Natural Resources and Life Sciences, Vienna, in cooperation with the Medical University of Vienna, since 2012 and are continuously adapted to reflect the latest developments in the field. The book addresses Master- and PhD students in biotechnology, molecular biology and immunology, and all those who are interested in antibody engineering.