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.  

Enzyme Technology is one the most promising disciplines in modern biotechnology. In this book, the applications of a wide variety of enzymes are highlighted. Current studies in enzyme technology are focused towards the discovery of novel enzymes (termed "bio-discovery" or "bio-prospecting") and the identification and elucidation of novel pathways of these novel enzymes with emphasis on their industrial relevance. With the development of molecular techniques and other bioinformatics tools, the time to integrate this subject with other fields in the life sciences has arrived. A rapid expansion of the knowledge base in the field of enzyme biotechnology has occurred over the past few years. Much of this expansion has been driven by the bio-discovery of many new enzymes from a wide range of environments, some extreme in nature, followed by subsequent protein (enzyme) engineering. These enzymes have found a wide range of applications, ranging from bioremediation, bio-monitoring, biosensor development, bioconversion to biofuels and other biotechnologically important value-added products. Hydrolases constitute a major component of the global annual revenue generated by industrial enzymes and the emphasis has therefore been placed on these enzymes and their applications. With the immense interest of researchers active in this area, this book will serve to provide information on current aspects in this field of study. In the current edition, the contributions of many diversified topics towards establishing new directions of research in the area of enzyme biotechnology are described. This book serves to provide a unique source of information to undergraduates, post graduates and doctoral courses in microbiology and biotechnology along with allied life sciences. The present edition of the book covers all important areas of enzyme biotechnology i.e. the wide variety of enzymes in the field of enzyme biotechnology and their industrial applications, new methods and state-of-the-art information on modern methods of enzyme discovery. This book will act as good resource on most of the current facets of enzyme technology for all students engaged in bioengineering and biotechnology.

Cell-free protein expression promises to narrow the technological gap between DNA and protein technologies and provide a platform for broad application of synthetic biology principles in the Life Sciences. It is a rapid and high throughput methodology for the conversion of DNA encoded genetic information into protein-mediated biochemical activities. Cell-Free Protein Synthesis: Methods and Protocols brings together the key opinion leaders of cell-free technology development and provides case studies and detailed protocols for the application of cell-free methodology. Chapters cover the main directions in the development of cell-free technologies including several recently developed cell-free systems, as well as a number of applications of cell-free systems ranging from discovery of biofuel enzymes to in vitro assembly of viruses. 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, Cell-Free Protein Synthesis: Methods and Protocols seeks to serve a wide variety of scientists with its well-honed methodologies.

In this thesis, the author outlines the development of new monolithic columns and isotope dimethyl labeling strategies and their applications in high-performance proteome analyses. Though different types of monolithic columns have been widely developed for chromatography and electrophoresis separation, their application in proteomics for complex peptide mixtures separation is still a challenge. The author discusses the preparation of new monolithic columns and optimization of chromatography separation capability to improve coverage and accuracy of proteome analysis. Further, the author describes a novel online multidimensional chromatography system combined with automated online isotope labeling, which significantly improves the throughput, sensitivity and accuracy of quantitative proteomics. In addition to the development of new technologies, the author investigates the proteome and phosphoproteome expression changes of clinical hepatocellular carcinoma tissues and the hippocampi of mice with Alzheimer’s disease. The work in this thesis has led to several publications in high-profile journals in the fields of analytical chemistry and proteome research.

Next Generation Sequencing: Chemistry, Technology and Applications, by P. Hui Application of Next Generation Sequencing to Molecular Diagnosis of Inherited Diseases, by W. Zhang, H. Cui, L.-J.C. Wong Clinical Applications of the Latest Molecular Diagnostics in Noninvasive Prenatal Diagnosis, by K.C.A. Chan The Role of Protein Structural Analysis in the Next Generation Sequencing Era, by W.W. Yue, D.S. Froese, P.E. Brennan Emerging Applications of Single-Cell Diagnostics, by M. Shirai, T. Taniguchi, H. Kambara Mass Spectrometry in High-Throughput Clinical Biomarker Assays: Multiple Reaction Monitoring, by C.E. Parker, D. Domanski, A.J. Percy, A.G. Chambers, A.G. Camenzind, D.S. Smith, C.H. Borchers Advances in MALDI Mass Spectrometry in Clinical Diagnostic Applications, by E.W.Y. Ng, M.Y.M. Wong, T.C.W. Poon Application of Mass Spectrometry in Newborn Screening: About Both Small Molecular Diseases and Lysosomal Storage Diseases, by W.-L. Hwu, Y.-H. Chien, N.-C. Lee, S.-F. Wang, S.-C. Chiang, L.-W. Hsu

To comprehend the organizational principle of cellular functions at diff erent levels, an integrative approach with large-scale experiments, the so-called 'omics' data including genomics, transcriptomics, proteomics, and metabolomics, is needed. Omics aims at the collective characterization and quantifi cation of pools of biological molecules that translate into the structure, function, and dynamics of an organism or organisms. Currently, omics is an essential tool to understand the molecular systems that underlie various plant functions. Furthermore, in several plant species, the development of omicsresources has progressed to address the particular biological properties of individual species. Integration of knowledge from omics-based research is an emerging issue as researchers seek to identify significance, gain biological insights and promote translational research. From these perspectives, we intend to provide the emerging aspects of plant systems research based on omics and bioinformatics analyses together with their associated resources and technological advances. Th e present book covers a wide range of omics topics, and discusses the latest trends and application area of plant sciences. In this volume, we have highlighted the working solutions as well as open problems and future challenges in plant omics studies. We believe that this book will initiate and introduce readers to state-of-the-art developments and trends in omics-driven research.

Featuring experimental approaches that shed light on the complexity of Ras GTPase biological functions, Ras Signaling: Methods and Protocols contains general overviews and detailed applications of both well-established and recently developed research techniques, including biochemical, biophysical, molecular biology, genetic and behavioral approaches, advanced high resolution fluorescence and electron microscopy imaging and "omics" technologies. Through this, the detailed volume provides information on expression, post-translational modifications, subcellular localization and dynamics, regulatory mechanisms of upstream and downstream signaling pathways and ultimately, biological activities and functions of Ras GTPases in different model systems, including high and low eukaryotic organisms. Written in the highly successful Methods in Molecular Biology series format, chapters include brief introductions, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols and tips on troubleshooting and avoiding known pitfalls. Wide-ranging and authoritative, Ras Signaling: Methods and Protocols serves as an aid for investigators of different backgrounds and interests related to the multiple physiological and pathological functions of the large superfamily of Ras GTPases.

The lipid-rich and otherwise challenging nature of many key tissues complicates many aspects of current research, and applications of the unique nature of lipoproteins and their biological effects has engendered unique and vital methodologies. In Lipoproteins and Cardiovascular Disease: Methods and Protocols, experts in the field present a compendium of advanced and classical molecular biology methods targeted towards lipoprotein, atherosclerosis, and vascular biology research, bringing together in a single volume an updated set of protocols and strategies for methods now driving the most recent advances, along with classical methods that are still widely used. Among the many topics covered in this cutting-edge work, the book delves into crucial techniques such as quantitative real-time PCR, microarrays, RT-PCR laser capture microdissection, and tissue-specific gene overexpression, knockout, and knockdown methodologies, including AAV as a liver-directed gene delivery vehicle. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective subjects, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and valuable notes which highlight tips on troubleshooting and avoiding known pitfalls. Comprehensive and easy to use, Lipoproteins and Cardiovascular Disease: Methods and Protocols serves both novices and experts alike as a complete guide for any researcher with an interest in lipoproteins and their significant biological effects.

This book is dedicated to the channels and pores that belong to an eclectic and ubiquitous class of unconventional - perhaps at times strange - pore-forming molecules, which nevertheless play fundamental roles in various organisms. These non-canonical channels may take on various and sometimes complex architectures, such as large beta-barrels or lipid-containing pores. They may originate from bacteria, viruses or intracellular organelles. For some of them, the physiologically relevant substrate may indeed be ions, and for others folded polypeptides. Some are released by cells in a soluble form that has the ability to insert into biological membranes to exert its permeabilizing effect. Many of these unconventional pores have been investigated by electrophysiology, which, by its virtue of focusing on a few or even a single unit, has provided invaluable insight into the mechanisms and structure-function relationships of these remarkable membrane entities. The chapters of this book highlight a representative set of these interesting investigations.

During the past decade as the data on gene sequences and expression patterns rapidly accumulated, cell-free protein synthesis technology has also experienced a revolution, becoming a powerful tool for the preparation of proteins for their functional and structural analysis. In Cell-Free Protein Production: Methods and Protocols, experts in the field contribute detailed techniques, the uses of which expand deep into the studies of biochemistry, molecular biology, and biotechnology. Beginning briefly with basic methods and historical aspects, the book continues with thorough coverage of protein preparation methods, the preparation of proteins that are generally difficult to prepare in their functional forms, applications of the cell-free technologies to protein engineering, as well as some methods that are expected to constitute a part of future technologies. Written in the highly successful Methods in Molecular Biology (TM) series format, the chapters include 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 cutting-edge, Cell-Free Protein Production: Methods and Protocols aims to help researchers continue the growth of the vital exploration of cell-free sciences and technologies in order to better understand the dynamic lives of cells.

This volume emphasizes new techniques to help understand protein cages and to apply them to a variety of technologies, highlighting the expertise of researchers based on three continents. Protein cages are currently inspiring diverse scientific disciplines and are therefore at the crossroads of extremely widely-scoped research, which is reflected in the detailed chapters of Protein Cages: Methods and Protocols. From nanomaterials studies and iron particles to computational strategies and Atomic Force Microscopy, the chapters herein collectively provide an introduction to the rich world of protein cage research and specific techniques to understand and exploit this fascinating class of proteins. Written in the highly successful Methods in Molecular Biology series format, chapters begin with an introduction to their respective topics, lists of the necessary materials, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Practical and cutting-edge, Protein Cages: Methods and Protocols will help to inspire and further propel the current multi-disciplinary enthusiasm in studying and discovering new applications for protein cages.

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.

The basic principle of electron crystallography is to calculate a 3D density map by combining the amplitudes obtained from electron diffraction patterns with the experimental phases calculated from images of two-dimensional crystals of membrane or soluble proteins. This technology is very well developed and has produced a number of atomic models of membrane proteins in a lipid environment. Focused on comprehensive experimental protocols, Electron Crystallography of Soluble and Membrane Proteins: Methods and Protocols covers the entire range of techniques used in electron crystallography, including protein sample preparation, 2D crystallization, and screening in negative stain over electron cryo-microscopy (cryo-EM) and data processing, as well as modeling of conformational changes. Additional chapters provide perspective on past, present, and future challenges as well as complementary methods. Written for the popular Methods in Molecular Biology (TM) series, the work contains the kind of detailed descriptions and implementation advice necessary to ensure successful results. Comprehensive and cutting-edge, Electron Crystallography of Soluble and Membrane Proteins: Methods and Protocols serves laboratories new to the methods as well as state-of-the-art facilities pursuing this exciting area of protein science.

How and why certain proteins misfold and how this misfolding is linked to many disease processes has become a well-documented topic of study. Protein Misfolding and Cellular Stress in Disease and Aging: Concepts and Protocols moves beyond the basics to emphasize the molecular effects of protein misfolding at a cellular level, to delineate the impacts and cellular reactions that play a role in pathogenetic mechanisms, and to pinpoint possible manipulations and treatment strategies that can counteract, modify, or delay the consequences of misfolding. The volume begins with several concepts and approaches developed in the recent past including a connection to the research field of aging, where protein misfolding diseases have been equated to premature aging processes, and the book's coverage continues with detailed descriptions of protocols for relevant experimental approaches. Written in the highly successful Methods in Molecular Biology (TM) series format, protocols 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, Protein Misfolding and Cellular Stress in Disease and Aging: Concepts and Protocols aims to aid researchers in the field, as well as medical professionals and molecular biologists, in shaping and performing research related to this intriguing and vital subject.

Expert authors provide critical, in-depth reviews of available methods for retrieving selective information out of complex biological systems. Sensors, probes and devices are present and future tools of medicinal diagnostics, environmental monitoring, food analysis and molecular biology. These are based on fluorescence, electrochemistry and mass spectrometry. Coverage of this volume includes sensor development for the detection of small analytes, monitoring of biomolecular interactions, analysis of cellular function, development of diagnostic tools.

This volume highlights recent developments in flow cytometry, affinity assays, imaging, mass spectrometry, microfluidics and other technologies that enable analysis of proteins at the single cell level. The book also includes chapters covering a suite of biochemical and biophysical methods capable of making an entire gamut of proteomic measurements, including analysis of protein abundance or expression, protein interaction networks, post-translational modifications, translocation and enzymatic activity. 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 thorough, Single Cell Protein Analysis: Methods and Protocols is useful to researchers and students in biological and biomedical sciences who have an interest in proteomic measurements in cells.

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 excellent book covers wide-ranging topics in interdisciplinary microbiology, addressing various research aspects and highlighting advanced discoveries and innovations. It presents the fascinating topic of modern biotechnology, including agricultural microbiology, microalgae biotechnology, bio-energy, bioinformatics and metagenomics, environmental microbiology, enzyme technology and marine biology. It presents the most up-to-date areas of microbiology with an emphasis on shedding light on biotechnological advancements and integrating these interdisciplinary microbiology research topics into other biotechnology sub-disciplines. The book raises awareness of the industrial relevance of microbiology, which is key component of this unique collection. The topics include production of antioxidant-glutathione, enzyme-engineering methods, probiotic microbiology and features of microbial xylanases. It also covers some other remarkable aspects of microbiology, like potential health hazards in recreational water and fullerene nanocomposites, which are vital for biotechnological interventions. This book will be valuable resource for senior undergraduate and graduate students, researchers and other interested professionals or groups working in the interdisciplinary areas of microbiology and biotechnology.

Immunology has made significant progress in the past decade, driven forward by rapidly advancing technology and a renewed interest in the vast realm of innate immunity. The receptors that mediate these functions are at the front lines of both protective and regulative roles of the immune system. In Immune Receptors: Methods and Protocols, expert researchers present a variety of experimental approaches to the characterization of immune receptors and the cell biology that mediates their functions. These include imaging techniques that aim to understand receptor localization and trafficking, techniques to measure receptor-ligand interactions, strategies to identify novel ligands and methods to analyze downstream receptor signaling, as well as strategies for genomic and proteomic characterization of receptor repertoires. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include 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. Authoritative and practical, Immune Receptors: Methods and Protocols offers technical descriptions and protocols that will be useful both to investigators who are interested in carrying out these procedures and to those who seek a deeper understanding of the bench science that lies behind the immunology literature.

Transcriptional regulation controls the basic processes of life. Its complex, dynamic, and hierarchical networks control the momentary availability of messenger RNAs for protein synthesis. Transcriptional regulation is key to cell division, development, tissue differen- ation, and cancer as discussed in Chapters 1 and 2. We have witnessed rapid, major developments at the intersection of computational biology, experimental technology, and statistics. A decade ago, researches were struggling with notoriously challenging predictions of isolated binding sites from low-throughput experiments. Now we can accurately predict cis-regulatory modules, conserved cl- ters of binding sites (Chapters 13 and 15), partly based on high-throughput ch- matin immunoprecipitation experiments in which tens of millions of DNA segments are sequenced by massively parallel, next-generation sequencers (ChIP-seq, Chapters 9, 10, and 11). These spectacular developments have allowed for the genome-wide mappings of tens of thousands of transcription factor binding sites in yeast, bacteria, mammals, insects, worms, and plants. Please also note the no less spectacular failures in many laboratories around the world.

The use of proteomics to study complex diseases such as cardiovascular disease, the leading cause of death in developed countries, has grown exponentially in recent years. Proteomics is a rapidly expanding investigation platform in cardiovascular medicine and is becoming integrated and incorporated into cardiovascular research. The proteomics field continues to develop with major improvements in mass spectrometry instrumentation, methodology and data analysis. Heart Proteomics: Methods and Protocols complies a selection of techniques and methods that target the numerous processes implicated in the pathophysiology of heart. Chapters cover protocols and updated methods in the heart proteomic area with a particular focus on MS-based methods of protein and peptide quantification and the analysis of posttranslational modifications as well as descriptions of system biology approaches, which provide a better understanding of normal and pathological processes. Written in the successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Heart Proteomics: Methods and Protocols is a representative selection of methods that will prove to be a useful resource for experienced proteomics practitioners and newcomers alike.

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!

Proteomics by means of mass spectrometry has rapidly changed the way that we analyze proteomes. Gel-Free Proteomics: Methods and Protocols addresses contemporary methods for gel-free proteome research with a special focus on differential analysis and protein modifications. Divided into twenty-five chapters, this detailed volume meticulously describes vital procedures needed to perform gel-free proteomics, ranging from sample preparation, isotope labeling for differential proteomics, enrichment technologies for modified proteins and peptides, and bioinformatics. Written in the successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Gel-Free Proteomics: Methods and Protocols serves as a timely resource for both professionals and novices pursing research in this critical field.

This second of two volumes discusses subfamily proteins which function in molecular and vesicular transport mechanisms inside the cell. In this volume the focus lies on the Rab, Ran and Arf subfamily members. As in Volume 1, the book is written by international renowned scientists in the field of small G-proteins. In elaborate reviews, biochemistry, structure, function and G-protein - effector interactions are described. Together with Volume 1 this book provides an comprehensive state-of-the-art work on small G-proteins (GTPases). It is written for Graduates and Professors in Biochemistry and Cell Biology interested in the mechanism and function of small G-proteins but are extremely valuable for those who want to move into the field.

Progress in functional proteomics has been limited for a long time, partially caused by limitations in assay sensitivity and sample capacity; however, protein microarrays have the ability to overcome these limitations so that a highly parallel analysis of hundreds of proteins in thousands of samples is attainable. In Protein Microarrays: Methods and Protocols, expert researchers in the field present an up-to-date collection of robust strategies in the field of protein microarrays and summarize recent advantages in the field of printing technologies, the development of suitable surface materials, as well as detection and quantification technologies. 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 notes on troubleshooting and avoiding known pitfalls. Comprehensive and cutting-edge, Protein Microarrays: Methods and Protocols aims to stimulate the application and further advancement of this powerful technology in labs worldwide.