Isolation and Purification of Proteins

Isolation and Purification of Proteins

This publication details the isolation of proteins from biological materials, techniques for solid-liquid separation, concentration, crystallization, chromatography, scale-up, process monitoring, product formulation, and regulatory and ...

Author: Rajni Hatti-Kaul

Publisher: CRC Press

ISBN: 0203911687

Category: Medical

Page: 676

View: 972

This publication details the isolation of proteins from biological materials, techniques for solid-liquid separation, concentration, crystallization, chromatography, scale-up, process monitoring, product formulation, and regulatory and commercial considerations in protein production. The authors discuss the release of protein from a biological host, selectivity in affinity chromatography, precipitation of proteins (both non-specific and specific), extraction for rapid protein isolation, adsorption as an initial step for the capture of proteins, scale-up and commercial production of recombinant proteins, and process monitoring in downstream processing.
Categories: Medical

Protein Purification Protocols

Protein Purification Protocols

Protein Purification Protocols provides all that is needed to design and carry out a successful purification program.

Author: Shawn Doonan

Publisher: Springer Science & Business Media

ISBN: 9781592595457

Category: Biochemistry

Page: 405

View: 947

A comprehensive collection of essential, time-tested recipes for successful protein fractionation and purification in any experimental circumstance. The protocols give step-by-step instructions on how to select a source for the protein of interest, how to obtain a usable initial extract, how to purify the protein from that extract using both chemical and molecular methods, and how to dry and store the purified protein. Protein Purification Protocols provides all that is needed to design and carry out a successful purification program. It helps both experienced and novice investigators to clarify and define their purification problems and then provides a comprehensive set of tools for a practical solution.
Categories: Biochemistry

Guide to Protein Purification

Guide to Protein Purification

Key Features * Detailed procedures newly written for this volume * Extensive practical information * Rationale and strategies for protein and enzyme purification * Personal perspectives on enzyme purification by eminent researchers Among ...

Author: Murray P. Deutscher

Publisher: Gulf Professional Publishing

ISBN: 0121820831

Category: Science

Page: 894

View: 340

Guide to Protein Purification, designed to serve the needs of the student, experienced researcher and newcomer to the field, is a comprehensive manual that provides all the up-to-date procedures necessary for purifying, characterizing, and handling proteins and enzymes in one source. Key Features * Detailed procedures newly written for this volume * Extensive practical information * Rationale and strategies for protein and enzyme purification * Personal perspectives on enzyme purification by eminent researchers Among the Topics Covered * General methods for handling proteins and enzymes * Extraction, subcellular fractionation, and solubilization procedures * Comprehensive purification techniques * Specialized purification procedures * Protein characterization * Immunological procedures * Computer analysis of protein structure
Categories: Science

Protein Purification and Its Application to Crystallization

Protein Purification and Its Application to Crystallization

Structural studies on proteins depend on an investigator 's ability to isolate and purify the protein.

Author: Jeffrey Robert Deschamps

Publisher:

ISBN: OCLC:21342811

Category: Crystallization

Page: 46

View: 360

Structural studies on proteins depend on an investigator 's ability to isolate and purify the protein. In many cases, protein isolation is a trivial matter, but purification to homogeneity is a lengthy process. Traditionally, proteins have been purified by a combination of precipitation, open-column chromatography, including size-exclusion and ion-exchange chromatography, ultracentrifugation, and electrophoresis. Recently, high-performance liquid chromatography(HPLC)has become increasingly important in the purification and analysis of proteins. The purpose of this study was to apply modern HPLC techniques to the protein purification problems encountered in crystallographic studies. (aw).
Categories: Crystallization

Cytoskeleton Proteins

Cytoskeleton Proteins

The standard protocols for the purification of all known cytoskeleton proteins are presented in this manual.

Author: Gerhard Isenberg

Publisher: Springer Science & Business Media

ISBN: 9783642796326

Category: Science

Page: 269

View: 909

The standard protocols for the purification of all known cytoskeleton proteins are presented in this manual. Proteins are listed alphabetically and each protocol follows a common format. Thus, the manual provides a quick and easy reference to all relevant procedures for cytoskeleton protein purification. The isolation procedure for each protein is shown in a clear flowchart, while the source of the protein, equipment and material needed, a list of suppliers, standard references, accession No. of sequences as well as further relevant facts and practical tips are given on a separate page.
Categories: Science

Purification and Analysis of Recombinant Proteins

Purification and Analysis of Recombinant Proteins

Covering both new and traditional topics in the purification and analysis of recombinant proteins, this volume demonstrates how to overcome problems in protein research and presents practical methods used in protein work, explaining their ...

Author: SEETHARAM

Publisher: CRC Press

ISBN: 0824782771

Category: Medical

Page: 344

View: 421

Covering both new and traditional topics in the purification and analysis of recombinant proteins, this volume demonstrates how to overcome problems in protein research and presents practical methods used in protein work, explaining their theoretical bases. The collection also explores innovative co
Categories: Medical

The Separation and Isolation of Proteins

The Separation and Isolation of Proteins

This volume will be of interest to chemists working with proteins.

Author: P. Alexander

Publisher: Elsevier

ISBN: 9781483149806

Category: Science

Page: 262

View: 182

A Laboratory Manual of Analytical Methods of Protein Chemistry (Including Polypeptides), Volume 1: The Separation and Isolation of Proteins deals with the techniques used in the separation and isolation of proteins, including fractionation and characterization by dialysis, multi-membrane electrodecantation, and zonal density gradient electrophoresis. The fractionation of proteins by adsorption and ion exchange is also described. This book is comprised of seven chapters and begins with a discussion on procedures for the separation of proteins, paying particular attention to the liberation of proteins from cellular material; removal of lipids from lipoproteins; and denaturation, fractionation, and purification of proteins. The next chapter focuses on the isolation of biologically active proteins such as cytochrome, bacterial amylases, and bacterial proteinases. The reader is methodically introduced to fractionation of proteins by adsorption and ion exchange; fractionation and characterization by dialysis; multi-membrane electrodecantation; and continuous and discontinuous partition. The final chapter explains how zonal density gradient electrophoresis works as a separation method for natural mixtures of proteins, their degradation products, and other substances carrying electric charges in solution or suspension. This volume will be of interest to chemists working with proteins.
Categories: Science

The Use of Molecular Fusion Endo tags for the Isolation and Purification of Multimeric Recombinant Proteins

The Use of Molecular Fusion Endo tags for the Isolation and Purification of Multimeric Recombinant Proteins

RESULTS: (1).

Author: Nihal Ramsis Yanni Magar

Publisher:

ISBN: OCLC:851561506

Category: DNA-binding proteins

Page: 520

View: 162

RESULTS: (1). The dimeric GFP was successfully expressed and purified, and the two GFP domains were separated by cleavage with rTEV protease. The endo-tagged dimeric GFP, and the C-terminally tagged GFP obtained after cleavage were each eluted from the Ni-NTA resin with the same concentration of imidazole, 100 mM. The GFP retained fluorescent activity, and the dimeric GFP showed approximately twice the fluorescence of the single GFP molecules. (2). The six plasmids incorporating gene cassettes encoding amylase S and xylanase in either order, with the three different endo-tags, were used to express the recombinant protein. Xylanase enzyme activity was measured in the clarified lysate from each preparation indicating that the recombinant multi-subunit proteins were expressed, but amylase enzyme activity was only detected from preparations containing construct I, amylase-xylanase, and not in construct 2, xylanase-amylase. For all six preparations, the level of expression of the full length protein was poor, and there was evidence of production of truncated proteins. None of the six preparations gave good binding and purification on Ni-NTA, and in all cases the proteins were progressively eluted from the column during washing with concentrations of imidazole less than 100 mM. However the preparations containing the His-X3-His tag bound better than the shorter tags, and it was possible to prepare enough purified amylase-xylanase complex for cleavage with rTEV protease. Proteins of the appropriate size were obtained following cleavage, although neither enzyme showed any activity. As expected, the xylanase was found in the unbound fraction, whereas the C-terminally tagged amylase remained bound to the Ni-NTA resin, and required elution with 1000 mM imidazole. CONCLUSION: The study of GFP showed that an endo-tag could be used successfully to purify multi-subunit proteins, and could be useful to reduce the cost of production of recombinant proteins if twice the amount of active protein could be obtained from the same fermentation reaction. However, the results of the coordinate expression of the two enzymes amylase S and xylanase A have provided important information about the factors that need to be considered in designing such systems, including the order of the subunit proteins in the complex, and the size of the endotag to be used. The results indicate that prior knowledge of the structural characteristics of the component molecules is essential.
Categories: DNA-binding proteins

Protein Biotechnology

Protein Biotechnology

In a predecessor to this book, emphasis was placed on the role of protein isolation in the food industry, because at the time this reflected the interests of most of the participants at the course.

Author: Felix Franks

Publisher: Springer Science & Business Media

ISBN: 9781592594382

Category: Science

Page: 592

View: 796

Proteins are the servants of life. They occur in all com- nent parts of living organisms and are staggering in their fu- tional variety, despite their chemical similarity. Even the simplest single-cell organism contains a thousand different p- teins, fulfilling a wide range of life-supporting roles. Additions to the total number of known proteins are being made on an increasing scale through the discovery of mutant strains or their production by genetic manipulation. The total international protein literature could fill a medi- sized building and is growing at an ever-increasing rate. The reader might be forgiven for asking whether yet another book on proteins, their properties, and functions can serve a useful purpose. An explanation of the origin of this book may serve as justification. The authors form the tutorial team for an int- sive postexperience course on protein characterization or- nized by the Center for Professional Advancement, East Brunswick, New Jersey, an educational foundation. The course was first mounted in Amsterdam in 1982 and has since been repeated several times, in both Amsterdam and the US, with participants from North America and most European countries. In a predecessor to this book, emphasis was placed on the role of protein isolation in the food industry, because at the time this reflected the interests of most of the participants at the course. Today, isolated proteins for food use are extracted from yeasts, fungal sources, legumes, oilseeds, cereals, and leaves.
Categories: Science

Membrane Protein Purification and Crystallization

Membrane Protein Purification and Crystallization

This second edition of Membrane Protein Purification and Crystallization, A Practical Guide is written for bench scientists working in the fields of biochemistry, biology, and proteomic research.

Author: Carola Hunte

Publisher: Elsevier

ISBN: 9780080536170

Category: Science

Page: 316

View: 305

This second edition of Membrane Protein Purification and Crystallization, A Practical Guide is written for bench scientists working in the fields of biochemistry, biology, and proteomic research. This guide presents isolation and crystallization techniques in a concise form, emphasizing the critical aspects unique to membrane proteins. It explains the principles of the methods and provides protocols of general use, permitting researchers and students new to this area to adapt these techniques to their particular needs. This edition is not only an update but is comprised mainly of new contributions. It is the first monograph compiling the essential approaches for membrane protein crystallization, and emphasizes recent progress in production and purification of recombinant membrane proteins. Provides general guidelines and strategies for isolation and crystallization of membrane proteins Gives detailed protocols that have wide application, and low specialized equipment needs Emphasizes recent progress in production and purification of recombinant membrane proteins, especially of histidine-tagged and other affinity-epitope-tagged proteins Summarizes recent developments of Blue-Native PAGE, a high resolution separation technique, which is independent of the use of recombinant techniques, and is especially suited for proteomic analyses of membrane protein complexes Gives detailed protocols for membrane protein crystallization, and describes the production and use of antibody fragments for high resolution crystallization Presents a comprehensive guide to 2D-crystallization of membrane proteins
Categories: Science

HPLC of Peptides and Proteins

HPLC of Peptides and Proteins

Hands-on experts from academia and industry comprehensively describe how to successfully perform all the critical HPLC techniques needed for the analysis of peptides and proteins.

Author: Marie-Isabel Aguilar

Publisher: Springer Science & Business Media

ISBN: 9781592597420

Category: Medical

Page: 413

View: 183

Hands-on experts from academia and industry comprehensively describe how to successfully perform all the critical HPLC techniques needed for the analysis of peptides and proteins. The methods range from commonly used techniques to those for capillary to large-scale preparative isolation. The authors have also presented a number of specific applications as case studies to illustrate the analytical approaches to a particular separation or assay challenge, with examples drawn from contemporary fields in biochemistry and biotechnology. Follow step-by-step instructions that ensure experimental success Develop your own separation and analytical protocols for peptide and protein analysis.
Categories: Medical

Expression Isolation and Purification of Human Zona Pellucida Protein 3

Expression  Isolation and Purification of Human Zona Pellucida Protein 3

Mammalian fertilization involves interactions of sperm surface receptors with the ligands of the zona pellucida, an extracellular matrix surrounding the ovulated oocytes.

Author: Ting-Fung Chi

Publisher:

ISBN: OCLC:42368126

Category: Fertilization in virto, Human

Page: 200

View: 704

Categories: Fertilization in virto, Human