Walker The Proteomics Protocols Handbook

2005
ISBN: 978-1-59259-890-8
Verlag: Humana Press
Format: PDF
Kopierschutz: 1 - PDF Watermark

E-Book, Englisch, Deutsch, 969 Seiten

Reihe: Springer Protocols Handbooks

Hands-on researchers describe in step-by-step detail 73 proven laboratory methods and bioinformatics tools essential for analysis of the proteome. These cutting-edge techniques address such important tasks as sample preparation, 2D-PAGE, gel staining, mass spectrometry, and post-translational modification. There are also readily reproducible methods for protein expression profiling, identifying protein-protein interactions, and protein chip technology, as well as a range of newly developed methodologies for determining the structure and function of a protein. The bioinformatics tools include those for analyzing 2D-GEL patterns, protein modeling, and protein identification. All laboratory-based protocols follow the successful Methods in Molecular Biology™ series format, each offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.

Written for: Biochemists, Proteomic scientists, protein researchers, molecular biologists

Autoren/Hrsg.

Walker, John M.

Inhaltsverzeichnis

1;Preface;5
2;Contents;6
3;Contributors;11
4;Extraction and Solubilization of Proteins for Proteomic Studies;17
5;Preparation of Bacterial Samples for 2-D PAGE;35
6;Preparation of Yeast Samples for 2-D PAGE;43
7;Preparation of Mammalian Tissue Samples for Two- Dimensional Electrophoresis;47
8;Differential Detergent Fractionation of Eukaryotic Cells;52
9;Serum or Plasma Sample Preparation for Two- Dimensional Gel Electrophoresis;64
10;Preparation of Plant Protein Samples for 2-D PAGE;70
11;Laser-Assisted Microdissection in Proteomic Analyses;73
12;Purification of Cellular and Organelle Populations by Fluorescence- Activated Cell Sorting for Proteome Analysis;81
13;Purification of Nucleoli From Lymphoma Cells and Solubilization of Nucleolar Proteins for 2- DE Separation;93
14;Prefractionation of Complex Protein Mixture for 2-D PAGE Using Reversed- Phase Liquid Chromatography;100
15;Fractionation of Complex Proteomes by Microscale Solution Isoelectrofocusing Using ZOOM Ù IEF Fractionators to Improve Protein Profiling;109
16;Large-Format 2-D Polyacrylamide Gel Electrophoresis;130
17;Analysis of Membrane Proteins by Two-Dimensional Gels;143
18;2-D PAGE of High-Molecular-Mass Proteins;155
19;Using Ultra-Zoom Gels for High-Resolution Two- Dimensional Polyacrylamide Gel Electrophoresis;160
20;NEpHGE and pI Strip Proteomic 2-D Gel Electrophoretic Mapping of Lipid- Rich Membranes;175
21;Silver Staining of 2-D Gels;184
22;Zn2+ Reverse Staining Technique;192
23;Multiplexed Proteomics Technology for the Fluorescence Detection of Phosphorylation and Protein Expression Levels Using Pro-Q® Diamond and SYPRO® Ruby Dyes;208
24;Sensitive Quantitative Fluorescence Detection of Proteins in Gels Using SYPRO® Ruby Protein Gel Stain;216
25;Ruby Protein Gel Stain;216
26;Rapid, Sensitive Detection of Proteins in Minigels With Fluorescent Dyes;222
27;Differential In-Gel Electrophoresis in a High-Throughput Environment;230
28;Statistical Analysis of 2-D Gel Patterns;245
29;2-DE Databases on the World Wide Web;265
30;Computer Analysis of 2-D Images;273
31;Comparing 2-D Electrophoretic Gels Across Internet Databases;284
32;Sample Cleanup by Solid-Phase Extraction/Pipet-Tip Chromatography;311
33;Protein Identification by In-Gel Digestion and Mass Spectrometric Analysis;314
34;Peptide Sequences of 2-D Gel-Separated Protein Spots by Nanoelectrospray Tandem Mass Spectrometry;318
35;Identification of Proteins by MALDI-TOF MS;322
36;Sequencing of Tryptic Peptides Using Chemically Assisted Fragmentation and MALDI- PSD;328
37;The In Situ Characterization of Membrane-Immobilized 2-D PAGE-Separated Proteins Using Ink-Jet Technology;344
38;Protein Identification by Peptide Mass Fingerprinting;358
39;Analysis of the Proteomes in Human Tissues by In-Gel Isoelectric Focusing and Mass Spectrometry;369
40;Liquid Chromatography Coupled to MS for Proteome Analysis;377
41;Quantitative Analysis of Proteomes and Subproteomes by Isotope- Coded Affinity Tag and Solid- Phase Glycoprotein Capture;386
42;Amino Acid-Coded Mass Tagging for Quantitative Profiling of Differentially Expressed Proteins and Modifications in Cells;394
43;Mass-Coded Abundance Tagging for Protein Identification and Relative Abundance Determination in Proteomic Experiments;407
44;Virtual 2-D Gel Electrophoresis by MALDI Mass Spectrometry;416
45;Identification of Posttranslational Modification by Mass Spectrometry;429
46;Approaches to the O-Glycoproteome;436
47;Identification of Protein Phosphorylation Sites by Mass Spectrometry;455
48;Quantitative Analysis of Protein Phosphorylation Status and Protein Kinase Activity on Microarrays Using Pro- Q Ù Diamond Dye Technology;463
49;New Challenges and Strategies for Multiple Sequence Alignment in the Proteomics Era;470
50;The Clustal Series of Programs for Multiple Sequence Alignment;488
51;FASTA Servers for Sequence Similarity Search;498
52;Protein Sequence Analysis and Domain Identification;521
53;Mammalian Genes and Evolutionary Genomics;536
54;Computational Identification of Related Proteins;547
55;Protein Identification and Analysis Tools on the ExPASy Server;563
56;Protein Sequence Databases;600
57;Characterization of Proteins;610
58;Computational Prediction of ProteinÒProtein Interactions;619
59;The Yeast Two-Hybrid System for Detecting Interacting Proteins;643
60;Antibody-Affinity Purification to Detect Interacting Proteins;673
61;Biomolecular Interaction Analysis Coupled With Mass Spectrometry to Detect Interacting Proteins;679
62;Assessment of AntibodyÒAntigen Interaction Using SELDI Technology;689
63;Protein and Peptide Microarray-Based Assay Technology;698
64;Production of Protein Microarrays Using Robotic Pin Printing Technologies;711
65;PCR-Directed Protein In Situ Arrays;722
66;Site-Specific Immobilization of Proteins in a Microarray;730
67;A Guide to Protein Interaction Databases;739
68;Deriving Function From Structure;786
69;Comparative Protein Structure Modeling;815
70;Classification of Protein Sequences and Structures;845
71;How to Use Protein 1-D Structure Predicted by PROFphd;859
72;Classification of Protein Folds;886
73;Protein Threading;903
74;High-Throughput Crystallography for Structural Proteomics;921
75;Automated High-Throughput Protein Crystallization;936
76;NMR-Based Structure Determination of Proteins in Solution;948