Protein Analysis and Purification: Benchtop Techniques

Forsideomslag
Springer Science & Business Media, 1. dec. 2013 - 520 sider
How one goes about analyzing proteins is a constantly evolving field that is no longer solely the domain of the protein biochemist. Investi gators from diverse disciplines find themselves with the unanticipated task of identifying and analyzing a protein and studying its physical properties and biochemical interactions. In most cases, the ultimate goal remains understanding the role(s) that the target protein is playing in cellular physiology. It was my intention that this manual would make the initial steps in the discovery process less time consuming and less intimidating. This book is not meant to be read from cover to cover. The expanded Table of Contents and the index should help locate what you are seeking. My aim was to provide practically oriented information that will assist the experimentalist in benchtop problem solving. The appendices are filled with diverse information gleaned from catalogs, handbooks, and manuals that are presented in a distilled fashion designed to save trips to the library and calls to technical service representatives. The user is encouraged to expand on the tables and charts to fit individual experimental situations. This second edition pays homage to the computer explosion and the various genome projects that have revolutionized how benchtop scientific research is performed. Bioinformatics and In silica science are here to stay. However, the second edition still includes recipes for preparing buffers and methods for lysing cells.
 

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Indhold

An Overview of This Manual
1
Protein Structure
8
Tracking the Target Protein
24
Labeling Proteins Present at the Plasma Membrane
30
PROTocol 3 8 Labeling Isolated Proteins with Chloramine T36
36
PROTocol 3 11 Immunoprecipitation
42
would like to thank Paula Callaghan for giving me the opportunity
43
Analysis of ProteinProtein Interactions
58
Protein Database Searches
244
Identifying and Analyzing Posttranslational
259
NGlycosylation
267
Neuraminidase NA
273
B Phosphorylation
278
PROTocol 920 Renaturation of Immobilized Kinases
287
PROTocol 929 Phosphoamino Acid Analysis of Phosphoproteins Isolated
298
Isoprenylation
305

Safety Considerations
79
How one goes about analyzing proteins is a constantly evolving field
80
PROTocol 4 12 Measuring the pH of the Gel Slices
87
Recovery of Proteins from the Gel
105
Getting Started with Protein Purification
118
Subcellular Markers
128
Manipulating Proteins in Solution
136
Changing the Buffer by Gel Filtration
142
Membrane Proteins
153
Transfer and Detection of Proteins
177
Enzymatic Detection Methods
199
Removing Unwanted Background Signal from Xray Film
205
Identification of the Target Protein
211
Cleavage of Proteins Transferred to PVDF or NC Membranes
217
Chemical Cleavage
219
Microsequencing from PVDF Membranes
225
Preparation of Proteins for MS Analysis
231
Protein Identification by MS
238
Triton X114
311
Chromatography
324
E Hydrophobic Interaction Chromatography HIC
356
F Affinity Chromatography
363
21 Purifying a Histidine Tagged Recombinant
380
Recombinant Protein Techniques
386
Microsomal Membranes
392
Affinity Tags
402
A Safety Considerations
433
Purification of Antibody Using Protein A Affinity Columns
439
Nucleic Acids
451
Centrifugation
465
Proteases and Proteolytic Enzyme Inhibitors
480
Radioactivity
490
K List of Suppliers Vendors Manufacturers
502
various genome projects that have revolutionized how benchtop
503
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