Shotgun proteomic experiments provide qualitative and quantitative analytical information from biological samples ranging in complexity from simple bacterial isolates to higher eukaryotes such as plants and humans and even to communities of microbial organisms. Improvements to instrument performance, sample preparation, and informatic tools are increasing the scope and volume of data that can be analyzed by mass spectrometry (MS). To accommodate for these advances, it is becoming increasingly essential to choose and/or create tools that can not only scale well but also those that make more informed decisions using additional features within the data. Incorporating novel and existing tools into a scalable, modular workflow not only provides more accurate, contextualized perspectives of processed data, but it also generates detailed, standardized outputs that can be used for future studies dedicated to mining general analytical or biological features, anomalies, and trends. This research developed cyber-infrastructure that would allow a user to seamlessly run multiple analyses, store the results, and share processed data with other users. The work represented in this dissertation demonstrates successful implementation of an enhanced bioinformatics workflow designed to analyze raw data directly generated from MS instruments and to create fully-annotated reports of qualitative and quantitative protein information for large-scale proteomics experiments. Answering these questions requires several points of engagement between informatics and analytical understanding of the underlying biochemistry of the system under observation. Deriving meaningful information from analytical data can be achieved through linking together the concerted efforts of more focused, logistical questions. This study focuses on the following aspects of proteomics experiments: spectra to peptide matching, peptide to protein mapping, and protein quantification and differential expression. The interaction and usability of these analyses and other existing tools are also described. By constructing a workflow that allows high-throughput processing of massive datasets, data collected within the past decade can be standardized and updated with the most recent analyses.

This volume is a compendium of cutting-edge protocols for quantitative proteomics, and presents the most significant methods used in the field today. The focus on mass spectrometry (MS) is integral. Attention is given to state-of-the-art techniques for the characterization of the phosphoproteome and tandem MS for detection of inborn errors of metabolism. This volume is an indispensable resource in the search for novel biomarkers.

This is an in-depth guide to the theory and practice of analyzing raw mass spectrometry (MS) data in proteomics. The volume outlines available bioinformatics programs, algorithms, and databases available for MS data analysis. General guidelines for data analysis using search engines such as Mascot, Xtandem, and VEMS are provided, with specific attention to identifying poor quality data and optimizing search parameters.

The field of proteomics has developed rapidly over the past decade nurturing the need for a detailed introduction to the various informatics topics that underpin the main liquid chromatography tandem mass spectrometry (LC-MS/MS) protocols used for protein identification and quantitation. Proteins are a key component of any biological system, and monitoring proteins using LC-MS/MS proteomics is becoming commonplace in a wide range of biological research areas. However, many researchers treat proteomics software tools as a black box, drawing conclusions from the output of such tools without considering the nuances and limitations of the algorithms on which such software is based. This book seeks to address this situation by bringing together world experts to provide clear explanations of the key algorithms, workflows and analysis frameworks, so that users of proteomics data can be confident that they are using appropriate tools in suitable ways.

A key experiment in biomedical research is monitoring the expression of different proteins in order to detect changes that occur in biological systems under different experimental conditions. The method that is most widely used is the Western blot analysis. While Western blot is a workhorse in laboratories studying protein expression and has several advantages, it also has a number of significant limitations. In particular, the method is semi-quantitative with limited dynamic range. Western blot focuses on a single protein per sample with only a small number of representative samples analyzed in an experiment. New quantitative tools have been needed for some time to at least supplement, & possibly replace, the Western blot. Mass spectrometric methods have begun to compete with Western blot for routine quantitative analyses of proteins. One of these methods is based on the tandem mass spectrometry technique of selected reaction monitoring (SRM), which is also called multiple reaction monitoring (MRM). Selected reaction monitoring is actually an older tandem mass spectrometry technique, first described in the late 70s, that is widely utilized in the quantitative analysis of small molecules like drugs & metabolites. The use of selected reaction monitoring for the quantitative analysis of proteins has a number of advantages. Most importantly, it is fundamentally quantitative with a wide dynamic range. The output of the analysis is a numerical result that can range over several orders of magnitude. Other advantages include sufficient specificity & sensitivity to detect low abundance proteins in complex mixtures. Finally, selected reaction monitoring can be multiplexed to allow the quantitative analysis of relatively large numbers of proteins in a single sample in a single experiment. This Brief will explain both the theoretical & experimental details of the selected reaction monitoring experiment as it is applied to proteins. ​

PROVIDES STRATEGIES AND CONCEPTS FOR UNDERSTANDING CHEMICAL PROTEOMICS, AND ANALYZING PROTEIN FUNCTIONS, MODIFICATIONS, AND INTERACTIONS—EMPHASIZING MASS SPECTROMETRY THROUGHOUT Covering mass spectrometry for chemical proteomics, this book helps readers understand analytical strategies behind protein functions, their modifications and interactions, and applications in drug discovery. It provides a basic overview and presents concepts in chemical proteomics through three angles: Strategies, Technical Advances, and Applications. Chapters cover those many technical advances and applications in drug discovery, from target identification to validation and potential treatments. The first section of Mass Spectrometry-Based Chemical Proteomics starts by reviewing basic methods and recent advances in mass spectrometry for proteomics, including shotgun proteomics, quantitative proteomics, and data analyses. The next section covers a variety of techniques and strategies coupling chemical probes to MS-based proteomics to provide functional insights into the proteome. In the last section, it focuses on using chemical strategies to study protein post-translational modifications and high-order structures. Summarizes chemical proteomics, up-to-date concepts, analysis, and target validation Covers fundamentals and strategies, including the profiling of enzyme activities and protein-drug interactions Explains technical advances in the field and describes on shotgun proteomics, quantitative proteomics, and corresponding methods of software and database usage for proteomics Includes a wide variety of applications in drug discovery, from kinase inhibitors and intracellular drug targets to the chemoproteomics analysis of natural products Addresses an important tool in small molecule drug discovery, appealing to both academia and the pharmaceutical industry Mass Spectrometry-Based Chemical Proteomics is an excellent source of information for readers in both academia and industry in a variety of fields, including pharmaceutical sciences, drug discovery, molecular biology, bioinformatics, and analytical sciences.

The definitive introduction to data analysis in quantitative proteomics This book provides all the necessary knowledge about mass spectrometry based proteomics methods and computational and statistical approaches to pursue the planning, design and analysis of quantitative proteomics experiments. The author’s carefully constructed approach allows readers to easily make the transition into the field of quantitative proteomics. Through detailed descriptions of wet-lab methods, computational approaches and statistical tools, this book covers the full scope of a quantitative experiment, allowing readers to acquire new knowledge as well as acting as a useful reference work for more advanced readers. Computational and Statistical Methods for Protein Quantification by Mass Spectrometry: Introduces the use of mass spectrometry in protein quantification and how the bioinformatics challenges in this field can be solved using statistical methods and various software programs. Is illustrated by a large number of figures and examples as well as numerous exercises. Provides both clear and rigorous descriptions of methods and approaches. Is thoroughly indexed and cross-referenced, combining the strengths of a text book with the utility of a reference work. Features detailed discussions of both wet-lab approaches and statistical and computational methods. With clear and thorough descriptions of the various methods and approaches, this book is accessible to biologists, informaticians, and statisticians alike and is aimed at readers across the academic spectrum, from advanced undergraduate students to post doctorates entering the field.

Mass spectrometry (MS) based proteomics and intact protein analyses are important tools for the structural study of proteins and provide powerful methods for solving biochemical puzzles involving proteins. The work described in this dissertation is aimed at the development of novel, efficient, and information rich strategies for protein structure and sequence analysis. The approaches developed have been applied to analytes ranging from proteolytic peptides to large non-covalent protein complexes.

The content of this volume is designed to reach a wide audience, including those involved with relevant technologies such as electrophoresis and mass spectrometry, to those interested in how proteomics can benefit research. A wide range of techniques are discussed, each specifically designed to address different needs in proteomic analysis. The concluding chapter discusses the important issue related to handling large amounts of data accumulated in proteomic studies. - Discusses proteomics in the postgenomic age - Includes various strategies for quantitative proteomics - Covers the role of MS in structural functional proteomics and proteomics in drug discovery and bioinformatics