Computational Methods for Mass Spectrometry Proteomics

Proteomics is the study of the subsets of proteins present in different parts of an organism and how they change with time and varying conditions. Mass spectrometry is the leading technology used in proteomics, and the field relies heavily on bioinformatics to process and analyze the acquired data.   Since recent years have seen tremendous developments in instrumentation and proteomics–related bioinformatics, there is clearly a need for a solid introduction to the crossroads where proteomics and bioinformatics meet.
Computational Methods for Mass Spectrometry Proteomics describes the different instruments and methodologies used in proteomics in a unified manner. The authors put an emphasis on the computational methods for the different phases of a proteomics analysis, but the underlying principles in protein chemistry and instrument technology are also described. The book is illustrated by a number of figures and examples, and contains exercises for the reader. Written in an accessible yet rigorous style, it is a valuable reference for both informaticians and biologists.
Computational Methods for Mass Spectrometry Proteomics is suited for advanced undergraduate and graduate students of bioinformatics and molecular biology with an interest in proteomics. It also provides a good introduction and reference source for researchers new to proteomics, and for people who come into more peripheral contact with the field.

Spis treści:
Preface.
Acknowledgements. 
1 Protein, Proteome, and Proteomics.
1.1 Primary goals for studying proteomes.
1.2 Defining the protein. 
1.2.1 Protein identity. 
1.2.2 Splice variants.
1.2.3 Allelic variants – polymorphisms. 
1.2.4 Posttranslational modifications.
1.2.5 Protein isoforms. 
1.3 Protein properties – attributes and values.  
1.3.1 The amino acid sequence.
1 .3.2 Molecular mass.
1.3.3 Isoelectric point.
1.3.4 Hydrophobicity. 
1.3.5 Amino acid composition.
1.4 Posttranslational modifications.
1.5 Protein sequence databases. 
1.5.1 UniProt KnowledgeBase (Swiss–Prot/TrEMBL, PIR).
1.5.2 The NCBI non–redundant database.  
1.5.3 The International Protein Index (IPI). 
1.5.4 Time–instability of sequence databases.  
1.6 Identification and characterization of proteins. 
1.6.1 Top–down and bottom–up proteomics. 
1.6.2 Protein digestion into peptides.
1.7 Two approaches for bottom–up protein analysis by mass spectrometry.
1.7.1 MS – Peptide mass fingerprinting.  
1.7.2 MS/MS – Tandem MS. 
1.7.3 Combination approaches. 
1.7.4 Reducing the search space.
1.8 Instrument calibration and measuring errors. 
1.8.1 Calibration.
1.8.2 Accuracy and precision. 
1.9 Exercises.
1.10 Bibliographic notes.
2 Protein Separation – 2D Gel Electrophoresis.
2.1 Separation on molecular mass – SDS–PAGE. 
2.1.1 Estimating the protein mass.
2.2 Separation on isoelectric point – IEF.
2.3 Separation on mass and isoelectric point, 2D.
2.3.1 Transferring the proteins from the first to the second  dimension.   
2.3.2 Visualizing the proteins after separation. 
2.3.3 Problems. 
2.3.4 Excising the proteins. 
2.4 2D SDS–PAGE for (complete) proteomics.  
2.4.1 Identifying the proteins.
2.4.2 Quantification. 
2.4.3 Programs for treating and comparing gels. 
2.4.4 Comparing results from different experiments – DIGE.
2.5 Exercises.
2.6 Bibliographic notes.
3 Protein Digestion.
3.1 Experimental digestion. 
3.1.1 Cleavage specificity. 
3.1.2 Trypsin.  
3.1.3 Chymotrypsin. 
3.1.4 Other considerations for the c hoice of a protease.
3.1.5 Random cleavage.
3.1.6 Chemical cleavage. 
3.1.7 In–gel digestion.  
3.2 In silico digestion.
3.3 Exercises.
3.4 Bibliographic notes.
4 Peptide Separation – HPLC.
4.1 High Pressure Liquid Chromatography – HPLC.
4.2 Stationary phases and separation modes. 
4.2.1 Reverse phase chromatography, RP. 
4.2.2 Strong cation exchange chromatography, SCX. 
4.2.3 Other types of chromatography for proteomics. 
4.2.4 Tandem HPLC.
4.3 Component migration and retention time. 
4.4 The shape of the peaks. 
4.4.1 The width.  
4.4.2 Asymmetry. 
4.4.3 Resolution.  
4.5 Chromatography used for protein identification. 
4.5.1 Theoretical calculation of the retention time for reverse phase chromatography. 
4.6 Chromatography used for quantification. 
4.7 Exercises.
4.8 Bibliographic notes. 
5 Fundamentals of Mass Spectrometry.
5.1 The principle of mass spectrometry.
5.2 Ionization sources.  
5.2.1 MALDI – Matrix Assisted Laser Desorption Ionization.
5.2.2 ESI – Electrospray Ionization.
5.2.3 Other ionization sources. 
5.3 Mass analyzers.  
5.4 Isotopic composition of peptides.
5.4.1 Estimating the charge.
5.5 Fractional masses.  
5.5.1 Estimating one or two peptides in a peak complex.
5.6 The raw data.
5.7 Mass resolution and resolving power.
5.7.1 Isotopic resolution.
5.8 Exercises.
5.9 Bibliographic notes. 
6 Mass Spectrometry – MALDI–TOF.
6.1 Time–of–flight analyzers and their resolution. 
6.1.1 Time–to–mass converter.
6.1.2 Producing spectra. 
6.1.3 Ionization statistics.
6.2 Constructing the peak list. 
6.2.1 Noise.
6.2.2 Baseline correction. 
6.2.3 Smoothing and noise reduction.
6.2.4 Peak detection.
6. 2.5 Example. 
6.2.6 Intensity normalization. 
6.2.7 Calibration.
6.3 Peak list preprocessing. 
6.3.1 Monoisotoping and deisotoping.
6.3.2 Removing spurious peaks.
6.4 Peak list format.  
6.5 Automation of MALDI–TOF–MS.
6.6 Exercises.
6.7 Bibliographic notes. 
7 Protein Identification and Characterization by MS.
7.1 The main search procedure.
7.1.1 The experimental data.
7.1.2 The database – the theoretical data.  
7.1.3 Other search parameters.
7.1.4 Organization of the database.
7.2 The peptide mass comparison. 
7.2.1 Reasons why experimental masses may not match.
7.3 Database search and recalibration.
7.3.1 The search program MSA (Mass Spectra Analyzer).
7.3.2 Aldente.   
7.4 Score calculation. 
7.4.1 Score components.
7.4.2 Scoring scheme examples.
7.4.3 Identification from a protein mixture. 
7.5 Statistical significance – the P–value.
7.5.1 A priori probability for k matches. 
7.5.2 Simulation for determining the P–value. 
7.5.3 A simple Mascot search. 
7.6 Characterization. 
7.7 Exercises.
7.8 Bibliographic notes.
8 Tandem MS or MS/MS Analysis.
8.1 Peptide fragments. 
8.2 Fragmentation techniques.
8.3 MS/MS spectrometers. 
8.3.1 Analyzers for MS/MS. 
8.4 Different types of analyzers. 
8.4.1 TOF/TOF. 
8.4.2 Triple quadrupole (Triple quad).
8.4.3 Ion trap (IT). 
8.4.4 Fourier Transform Ion Cyclotron Resonance (FT–ICR).
8.4.5 Combining quadrupole and Time of flight – Q–TOF.
8.4.6 Combining quadrupole and ion trap – Q–TRAP.
8.4.7 Combining TOF and Ion trap.
8.4.8 Combining Linear ion trap with Orbitrap.
8.4.9 Characteristics and performances of some type of analyzers.
8.5 Overview of the process for MS/MS analysis.
8.6 Fragment ion masses and r