This book focuses on recent developments of Pichia pastoris as a recombinant protein production system. Highlighted topics include a discussion on the use of fermentors to grow Pichia pastoris, information on the O- and N-linked glycosylation, methods for labeling Pichia pastoris expressed proteins for structural studies, and the introduction of mutations in Pichia pastoris genes by the methods of restriction enzyme-mediated integration (REMI). Each chapter presents cutting-edge and cornerstone protocols for utilizing P. pastoris as a model recomibinant protein production system. This volume fully updates and expands upon the first edition.
Two of the recent books in the Methods in Molecular Biology series, Yeast Protocols and Pichia Protocols, have been narrowly focused on yeasts and, in the latter case, particular species of yeasts. Food Microbiology Pro- cols, of necessity, covers a very wide range of microorganisms. Our book treats four categories of microorganisms affecting foods: (1) Spoilage organisms; (2) pathogens; (3) microorganisms in fermented foods; and (4) microorganisms p- ducing metabolites that affect the flavor or nutritive value of foods. Detailed information is given on each of these categories. There are several chapters devoted to the microorganisms associated with fermented foods: these are of increasing importance in food microbiology, and include one bacteriophage that kills the lactic acid bacteria involved in the manufacture of different foods—cottage cheese, yogurt, sauerkraut, and many others. The other nine chapters give procedures for the maintenance of lactic acid bacteria, the isolation of plasmid and genomic DNA from species of Lac- bacillus, determination of the proteolytic activity of lactic acid bacteria, det- mination of bacteriocins, and other important topics.
This volume provides an overview of the main yeast production platforms currently used and future yeast cell factories for recombinant protein production. Chapters detail approaches of genetic and metabolic engineering, co-factor containing proteins and virus-like particles, glycoproteins, and post-translational modifications of proteins. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Recombinant Protein Production in Yeast: Methods and Protocols aims to provide state of the art background and methods for protein producing yeast platforms, as well as case studies for special applications.
While the choices of microbial and eukaryotic expression systems for production of recombinant proteins are many, most researchers in academic and industrial settings do not have ready access to pertinent biological and technical information since it is normally scattered throughout the scientific literature. This book closes the gap by providing information on the general biology of the host organism, a description of the expression platform, a methodological section -- with strains, genetic elements, vectors and special methods, where applicable -- as well as examples of proteins produced with the respective platform. The systems thus described are well balanced by the inclusion of three prokaryotes (two Gram-negatives and one Gram-positive), four yeasts, two filamentous fungi and two higher eukaryotic cell systems -- mammalian and plant cells. Throughout, the book provides valuable practical and theoretical information on the criteria and schemes for selecting the appropriate expression platform, the possibility and practicality of a universal expression vector, and on comparative industrial-scale fermentation, with the production of a recombinant Hepatitis B vaccine chosen as an industrial example. With a foreword by Herbert P. Schweizer, Colorado State University, USA: "As a whole, this book is a valuable and overdue resource for a varied audience. It is a practical guide for academic and industrial researchers who are confronted with the design of the most suitable expression platform for their favorite protein for technical or pharmaceutical purposes. In addition, the book is also a valuable study resource for professors and students in the fields of applied biology and biotechnology."
I belie ve that the book would provide an overview of the recent developments in the domain of yeast research with some new ideas, which could serve as an inspiration and challenge for researchers in this field. Ne w Delhi Prof. Asis Datta Dec. 24, 2007 F ormer Vice-chancellor, JNU Director, NCPGR (New Delhi) Pr eface Yeasts are eukaryotic unicellular microfungi that are widely distributed in the natural environments. Although yeasts are not as ubiquitous as bacteria in the na- ral environments, they have been isolated from terrestrial, aquatic and atmospheric environments. Yeast communities have been found in association with plants, a- mals and insects. Several species of yeasts have also been isolated from specialized or extreme environments like those with low water potential (e. g. high sugar/salt concentrations), low temperature (e. g. yeasts isolated from Antarctica), and low oxygen availability (e. g. intestinal tracts of animals). Around 1500 species of yeasts belonging to over 100 genera have been described so far. It is estimated that only 1% of the extant yeasts on earth have been described till date. Therefore, global efforts are underway to recover new yeast species from a variety of normal and extreme environments. Yeasts play an important role in food chains, and carbon, nitrogen and sulphur cycles. Yeasts can be genetically manipulated by hybridization, mutation, rare m- ing, cytoduction, spheroplast fusion, single chromosomal transfer and transfor- tion using recombinant technology. Yeasts (e. g.
This book reviews preparation of expression vectors, generation of high-yielding clones, scale-up, disruption of yeast cells to enable isolation of recombinant protein prior to purification and more, in the popular Methods in Molecular Biology format."
Yeast Metabolic Engineering: Methods and Protocols provides the widely established basic tools used in yeast metabolic engineering, while describing in deeper detail novel and innovative methods that have valuable potential to improve metabolic engineering strategies in industrial biotechnology applications. Beginning with an extensive section on molecular tools and technology for yeast engineering, this detailed volume is not limited to methods for Saccharomyces cerevisiae, but describes tools and protocols for engineering other yeasts of biotechnological interest, such as Pichia pastoris, Hansenula polymorpha and Zygosaccharomyces bailii. Tools and technologies for the investigation and determination of yeast metabolic features are described in detail as well as metabolic models and their application for yeast metabolic engineering, while a chapter describing patenting and regulations with a special glance at yeast biotechnology closes the volume. Written in the highly successful Methods in Molecular Biology series format, most chapters include an introduction to their respective topic, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols and tips on troubleshooting and avoiding known pitfalls. Comprehensive and authoritative, Yeast Metabolic Engineering: Methods and Protocols aims to familiarize researchers with the current state of these vital and increasingly useful technologies.
The general field of fundamental and applied biotechnology becomes increasingly important for the production of biologicals for human and veterinary use, by using prokaryotic and eukaryotic microorganisms. The papers in the present book are refereed articles compiled from oral and poster presentations from the EFB Meeting on Recombinant Protein Production with Prokaryotic and Eukaryotic Cells. A Comparative View on Host Physiology, which was organized in Semmering/A from 5th to 8th October 2000. A special feature of this meeting was the comparison of different classes of host cells, mainly bacteria, yeasts, filamentous fungi, and animal cells, which made obvious that many physiological features of recombinant protein formation, like cell nutrition, stress responses, protein folding and secretion, or genetic stability, follow similar patterns in different expression systems. This comparative aspect is by far the point of most interest because such comparisons are rarely done, and if they are done, their results are most often kept secret by the companies who generated them. Audience: Presently, a comparable book does not exist because the compiling of manuscripts from all fields of biotechnology (prokaryotic as well as eukaryotic, up to animal cell biotechnology) is not done in general. This particularity makes this book very interesting for postgraduate students and professionals in the large field of biotechnology who want to get a more global view on the current state of the expression of recombinant biologicals in different host cell systems, the physiological problems associated with the use of different expression systems, potential approaches to solve such difficulties by metabolic engineering or the use of other host cells, and the cooperation between process development and strain improvement, which is crucial for the optimisation of both the production strain and the process. This book should be in every library of an institution/organization involved in biotechnology.
Designed as a research-level guide to current strategies and methods of membrane protein production on the small to intermediate scale, this practice-oriented book provides detailed, step-by-step laboratory protocols as well as an explanation of the principles behind each method, together with a discussion of its relative advantages and disadvantages. Following an introductory section on current challenges in membrane protein production, the book goes on to look at expression systems, emerging methods and approaches, and protein specific considerations. Case studies illustrate how to select or sample the optimal production system for any desired membrane protein, saving both time and money on the laboratory as well as the technical production scale. Unique in its coverage of "difficult" proteins with large membrane-embedded domains, proteins from extremophiles, peripheral membrane proteins, and protein fragments.
Laboratory Protocols in Fungal Biology presents the latest techniques in fungal biology. This book analyzes information derived through real experiments, and focuses on cutting edge techniques in the field. The book comprises 57 chapters contributed from internationally recognised scientists and researchers. Experts in the field have provided up-to-date protocols covering a range of frequently used methods in fungal biology. Almost all important methods available in the area of fungal biology viz. taxonomic keys in fungi; histopathological and microscopy techniques; proteomics methods; genomics methods; industrial applications and related techniques; and bioinformatics tools in fungi are covered and complied in one book. Chapters include introductions to their respective topics, list of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and notes on troubleshooting. Each chapter is self-contained and written in a style that enables the reader to progress from elementary concepts to advanced research techniques. Laboratory Protocols in Fungal Biology is a valuable tool for both beginner research workers and experienced professionals. Coming Soon in the Fungal Biology series: Goyal, Manoharachary / Future Challenges in Crop Protection Against Fungal Pathogens Martín, García-Estrada, Zeilinger / Biosynthesis and Molecular Genetics of Fungal Secondary Metabolites Zeilinger, Martín, García-Estrada / Biosynthesis and Molecular Genetics of Fungal Secondary Metabolites, Volume 2 van den Berg, Maruthachalam / Genetic Transformation Systems in Fungi Schmoll, Dattenbock / Gene Expression Systems in Fungi Dahms / Advanced Microscopy in Mycology
