Preface
Editor
List of contributing authors
Abbreviations
Acknowledgements
1 Three-phase partitioning
1.1 Method
1.2 The mechanism of TPP
1.3 A practical example β the isolation of cathepsin L from liver tissue
1.4 Other applications
2 Folding and degradation functions of molecular chaperones
2.1 Introduction
2.2 The domain structure of Hsc/Hsp70
2.3 The Hsc/Hsp70 reaction cycle
2.4 Cochaperones determine the function of Hsc/Hsp70
2.5 In vitro reconstitution and functional analysis of the Hsc/Hsp70 chaperone system
2.6 Measuring the ATPase activity of Hsc/Hsp70
2.7 Determining chaperone activity
2.8 In vitro reconstitution of chaperone-assisted ubiquitylation
2.9 Concluding remarks
3 Membrane protein folding in detergents
3.1 Introduction
3.2 Interactions of membrane proteins with detergents
3.3 Techniques to characterize TM proteins in detergents
3.4 Applications of TM protein-detergent complexes
3.5 Conclusions
4 Glycoprotein-folding quality control in the endoplasmic reticulum
4.1 Introduction
4.2 Glycoprotein-folding quality control (QC)
4.3 The UGGT
4.4 GII
4.5 CNX and CRT
4.6 ERp57
4.7 Methods to study glycoprotein folding QC
5 Conformational dynamics in peptides and proteins studied by triplet-triplet energy transfer
5.1 Introduction
5.2 Concept of TTET experiments to study intrachain loop formation in polypeptide chains
5.3 Diffusion-controlled loop formation in unstructured polypeptide chains
5.4 Detection of fast conformational fluctuations in folded peptides and proteins by TTET
5.5 Conclusions
6 Protein import into the intermembrane space of mitochondria
6.1 Introduction
6.2 The mitochondrial IMS
6.3 The mitochondrial disulfide relay
6.4 The sulfhydryl oxidase Erv1
6.5 The oxidoreductase Mia40
6.6 Substrates of the mitochondrial disulfide relay
6.7 Methods to study mitochondrial protein translocation
6.8 General comments to the analysis of thiol-disulfide redox states
6.9 Outlook
7 On-membrane identification of gel-resolved proteins by matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS)
7.1 Introduction
7.2 Methods for identifying proteins electroblotted onto the PVDF membrane
7.3 General comments to the analysis of proteins on membranes
7.4 PVDF membranes or diamond-like carbon-coated (DLC) stainless steel plates?
7.5 Concluding remarks
8 Analysis of protein complexes using chemical cross-linking and mass spectrometry
8.1 Introduction
8.2 Reagents for chemical cross-linking
8.3 The chemical cross-linking workflow
8.4 MS and data analysis
8.5 Practical examples
8.6 The use of spatial constraints for modeling
8.7 Conclusion and outlook
9 Single-crystal spectroscopy correlated with X-ray crystallography provides complementary perspectives on macromolecular function
9.1 Introduction
9.2 Ionizing radiation: essential for crystal structures; a problem and a reagent
9.3 Cofactors in biology provide spectroscopic access to reaction cycles
9.4 Single-crystal spectroscopy correlated with X-ray diffraction
9.5 Correlated studies at beamline X26-C of the NSLS
9.6 Future prospects
10 Wide-angle X-ray solution scattering (WAXS)
10.1 Introduction
10.2 Sample preparation
10.3 Sample-handling robot
10.4 Data collection
10.5 Data processing
10.6 Structural information
10.7 Size and shape
10.8 Secondary and tertiary structure
10.9 Quaternary structure
10.10 Structural changes
10.11 Unfolding
10.12 Molecular modeling
10.13 Modeling of structural fluctuations
10.14 Outlook
11 Where purity matters: recombinant versus synthetic peptides in beta amyloid formation
11.1 Amyloid fibrils possess a defined quaternary structure
11.2 The importance of purity for reproducible kinetics of amyloid fibril formation in vitro: the AΓ as an example
11.3 Future challenges for the characterization of fibrillar structures
12 Chemical modification of proteins in living cells
12.1 Introduction
12.2 Site-specific labeling of proteins with chemical probes
12.3 Selecting an appropriate labeling technique
12.4 Live cell applications
12.5 Technical Protocols for SNAP- tag labeling
13 Proteomics of human bronchoalveolar lavage fluid: discovery of biomarkers of chronic obstructive pulmonary disease (COPD) with difference gel electrophoresis (DIGE) and mass spectrometry (MS)
13.1 Introduction
13.2 Application of DIGE platform to COPD biomarker discovery
13.3 Outlook
14 Proteomic analysis of Duchenne muscular dystrophy (DMD)
14.1 Introduction
14.2 Materials
14.3 Methods
14.4 Results and Discussion
14.5 Conclusion
15 Target-oriented peptide arrays in a palliative approach to cystic fibrosis (CF)
15.1 Introduction
15.2 PDZ domains
15.3 CF
15.4 Role of PDZ domains in CFTR trafficking
15.5 Target-oriented peptide arrays
15.6 An engineered peptide inhibitor of CAL extends the half-life of AF508-CFTR
15.7 Methods
15.8 Outlook
16 Probing protein dynamics in vivo using backbone cyclization: bacterial acyl carrier protein as a case study
16.1 Introduction
16.2 In vivo protein cyclization, biophysical analyses and functional assays
16.3 Outlook
17 The protein epitope mimetic approach to protein-protein interaction inhibitors
17.1 Introduction
17.2 Mechanisms of protein-protein interactions
17.3 Small-molecule screening approaches
17.4 Protein epitope mimetic approaches
18 The structural biology of Ξ±1-antitrypsin deficiency and the serpinopathies
18.1 Clinical phenotypes of the serpinopathies
18.2 The serpin mechanism of protease inhibition
18.3 Folding, misfolding and polymerization
18.4 Serpin folding
18.5 Dissecting the pathways of polymerization
18.6 Cellular processing of polymers
18.7 Stem cell technology to generate models of disease
18.8 Conclusions
Index