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In biomedical research, understanding and defining the origin of diseases and their effect on multiple organs is a necessity. It has to be done at several levels: genomic, transcriptomic, proteomic and metabolomic. When the origin of a disease is suspected to be monogenic, the best approach to unravel its cause is obviously genomic. When the behaviour or the aggression of a cancer and its response to therapy are related to multiple gene modifications or expression deregulation, tissue biopsies should be analysed at transcriptomic levels by reverse transcription PCR and DNA microarrays. When the disease impacts mostly on the internal environment and is due to the accumulation of ‘toxic' material affecting multiple organs, proteomic and metabolomic approaches are required. The present chapter discuss how proteomics can be used in medicine for basic or applied research, in fundamental or clinical domains, to unravel disease processes or to discover biomarkers and therapeutic targets. [ABSTRACT FROM AUTHOR]

In this chapter we review the changes to proteomics over the last decade and suggest where the field will go over the next 10 years. In the early days of proteomics, the focus was on large-scale protein identification either in a whole organisms or tissue. Changes in protein expression were often compared with emergence of disease etc. Today, the focus of basic research is moving towards protein characterisation (with emphasis on post-translational modifications) and protein-protein interaction mapping, with new informatics needed. Increasingly this leads to studies on function and towards practical applications of new discoveries. We suggest that proteomics is now close to providing the markers for new-generation diagnostic and prognostic tests, as well as the basis for finding new-generation drugs. [ABSTRACT FROM AUTHOR]

Protein-protein interactions are central to postgenome biology. Complex cellular functions are almost always the result of the coordinated action of several proteins, acting in molecular assemblies or pathways to achieve a particular task. The wiring schemes provided by protein interaction networks are believed to contribute to our understanding of the convoluted relationships between genomes and phenotypes. In human, impairment of pathway flow or deregulated connections can lead to abnormalities. The majority of targets of current therapeutics cluster in a limited number of pathways. One of the goals of this chapter is to exemplify and illustrate the importance of protein-protein interactions for human biology and pathology: how protein interaction networks contribute to the understanding of phenotype and more particularly human genetic disorders, helping the gene-finding process and bringing a molecular framework to the genetic heterogeneity and pleiotropy often observed in human syndromes. An important section of this chapter is also dedicated to a review of the proteomic technologies developed and adapted to chart protein-protein interactions on a whole cell or organism scale. Their respective advantages, shortcomings and limitations are presented. Finally, recent successes in the development of small molecules that target and interfere with protein-protein interaction sites open new avenues towards the design of drugs that specifically abrogate or modulate diseaserelevant interactions. This constitutes the last topic of the chapter. [ABSTRACT FROM AUTHOR]

The present chapter attempts to cover the recent initiatives directed towards representing, displaying and processing protein-related data suitable for interpreting proteomics data. Examples of integrated proteomics data are described and commented on. Challenging issues of integration are briefly discussed as well as directions for ongoing and future work. In particular, we consider the overlap between data integration, integrative biology and systems biology. [ABSTRACT FROM AUTHOR]

The field of proteomics has benefited enormously from the use of images, in particular those resulting from the analysis of samples by two-dimensional electrophoresis. Proteome imaging has the same function as a microscope: it allows the resolution of features and constituents of an object, in this case a proteome, which are otherwise invisible to the naked eye. Functionally, proteome imaging also allows differences between proteomic samples to be highlighted. The four major areas of proteome imaging, related to the underlying analytical proteomic technologies, are discussed in this chapter: two-dimensional electrophoresis gel images, images of the combination of liquid chromatography and mass spectrometry, images from the ‘molecular scanner' and mass spectrometry tissue images. Their common goal is to display proteomes in a form that is amenable to human vision and computer analysis, which can subsequently facilitate the comparison of two or more samples and assist protein identification. The background of the imaging procedures both in terms of the experimental requirements and the available software and tools is described, as well as the applications and use of proteome imaging for real proteomic studies. [ABSTRACT FROM AUTHOR]

The release of the sequence of the human genome in early 2000 generated enormous excitement with the promise of rapid identification of the gene products responsible for cellular function. What has become apparent is that knowledge of the DNA sequence of the gene that is involved in a particular metabolic process is insufficient to predict its function, expression and activity. The very numbers illustrate this difference - it is estimated that there are 22,000 to 25,000 known genes in the human genome but there are probably greater than 1,000,000 proteins in the human proteome. This discrepancy has extended the focus of proteomics to the analysis and understanding of the modifications that occur to proteins both during and after translation of the gene. As we move further into understanding protein function it is becoming increasingly obvious that many of the changes associated with disease and differentiation are to do with the modifications to the proteins rather than only to do with the regulation of the expression of the gene. The main difficulty which has slowed the understanding of the biological role of these protein modifications has been the perception that the analysis of these alterations is difficult and is best left to the limited number of experts in each field. However, the reality is that the increasing availability of sample preparation, mass spectrometric and bioinformatic tools specifically designed for the analysis of post-translational modifications, is enabling the function of these instruments of biological diversity to be explored. [ABSTRACT FROM AUTHOR]

The large-scale systematic quantitation of proteins is an important component of proteomics and has contributed to the emergence of the new fields of systems biology and molecular medicine. Quantitative proteomics is anticipated to provide new insights into biological function, facilitate the identification of diagnostic or prognostic disease markers, and contribute to the discovery of proteins as therapeutic targets. This field is growing rapidly and new applications and protocol design are driving technological innovations. Due to rapidly evolving innovations the field of quantitation is wide, and deep! So many topics cannot be discussed in such a short chapter. We advise the reader to gain an understanding of the computational procedures required for the processing of quantitative MS data, and ideally information on the various types of instruments that are best used. In this section we will provide a modest overview of the types of strategies employed in proteomics that rely on mass spectrometry for protein identification and concurrent quantitation. Throughout this review, comparative analysis is a recurring theme and one which is common to most quantitative techniques in proteomics. [ABSTRACT FROM AUTHOR]

Identifying proteins contained in a biological sample represents a central task in a majority of proteomics projects. The present chapter discusses various attributes that can be used to identify proteins: the species of origin, the isoelectric point, the amino acid composition, sequence tags and mass values. Then, the chapter focuses on protein identification using mass spectrometry data. It introduces the mass spectrometry technology, presents specific instrumentations and describes the major identification approaches, including peptide mass fingerprinting, peptide fragment fingerprinting and de novo sequencing. Available software tools are described, and for each approach, a concrete example of identification results is given. Particular care is attributed to the identification of proteins carrying post-translational modifications. [ABSTRACT FROM AUTHOR]

In the 1990s much of the technology development for 2-D gels was centred on attempts to solubilise and separate whole proteomes. In the last 10 years, much has changed in the science of sample preparation, but the art of 2-D gels is almost the same, and remains a method that many love to hate. The improvements in protein solubilisation, the introduction of pH gradients that extend into the far alkaline region, and particularly the hunt for lowabundance proteins have served to hasten the arrival of the most important current area of sample preparation - fractionation. As proteomics matures, there is a new focus on function and the analysis of co- and post-translational modifications. With this in mind, and as the complexity of sample preparations increases, this chapter deals with possible artefacts and how to avoid generating them. Finally, with artefacts banished, this chapter reviews the most appealing prefractionation tools that are currently available. Fractionation is discussed as a formidable tool for mining below the tip of the proteomic iceberg. [ABSTRACT FROM AUTHOR]

The concept of the proteome is now over 10 years old. As with all anniversaries, it is a good time to look back and reflect on what has been achieved in the area that we now call proteomics. What has been done well? What has been done not-so-well? What has been achieved, and what still eludes us? This review will briefly explore some of these questions, with respect to protein separations, mass spectrometry, and proteomic bioinformatics. [ABSTRACT FROM AUTHOR]

With the development of the Internet, a growing number of proteomics databases have become available. The web is a powerful tool for data integration because it links the components constituting these databases, in general gel images and protein information, while offering rapid means to navigate from one database to another. Unfortunately, with only 15 maps available as electronic resources, the human body fluids do not really benefit from this development. This chapter summarizes the state of the art of proteomics databases, with an emphasis on human body fluids. Insights into one of these databases, SWISS-2DPAGE, available for more than 10 yr now, are given to show current functionalities and usage examples. Some general thoughts are also given on how to improve sharing and publication of proteomics data through electronic media. [ABSTRACT FROM AUTHOR]

Protein identification and analysis software performs a central role in the investigation of proteins from two-dimensional (2-D) gels and mass spectrometry. For protein identification, the user matches certain empirically acquired information against a protein database to define a protein as already known or as novel. For protein analysis, information in protein databases can be used to predict certain properties about a protein, which can be useful for its empirical investigation. The two processes are thus complementary. Although there are numerous programs available for those applications, we have developed a set of original tools with a few main goals in mind. Specifically, these are: 1.To utilize the extensive annotation available in the Swiss-Prot database (1) wherever possible, in particular the position-specific annotation in the Swiss-Prot feature tables to take into account posttranslational modifications and protein processing.2.To develop tools specifically, but not exclusively, applicable to proteins prepared by twodimensional gel electrophoresis and peptide mass fingerprinting experiments.3.To make all tools available on the World-Wide Web (WWW), and freely usable by the scientific community. [ABSTRACT FROM AUTHOR]

Indeed, the development of two-dimensional (2-D) electrophoresis technology has provided an important research tool for comparative proteomics. A number of other technologies for protein separation have also been investigated in the last few years, such as protein chips, liquid chromatography, and high-pressure liquid chromatography, to name just a few. However, 2-D gels are still considered the workhorse among research laboratories willing to accurately track changes in protein expression and to achieve differential proteomics. [ABSTRACT FROM AUTHOR]

With the development of the Internet, a growing number of two-dimensional gel electrophoresis (2-DE) databases become available (50 in January 2004, for a total of 270 image maps). By linking the two components constituting 2-DE databases—images and protein information—the active hypertext links provide a powerful tool for data integration, in addition to navigation from one database to another. This chapter shows how to remotely retrieve data from these databases over the Internet. [ABSTRACT FROM AUTHOR]

Protein identification and analysis software performs a central role in the investigation of proteins from two-dimensional (2-D) gels. For protein identification, the user matches certain empirically gained information against a protein database to define a protein as already known or as novel. For protein analysis, information in protein databases can be used to predict certain properties about a protein, which can be useful for its empirical investigation. The two processes are thus complementary. Although there are numerable programs available for the above-mentioned applications, we have developed a set of original tools with a few main goals in mind. Specifically, these are: 1.To utilize the extensive annotation available in the SWISS-PROT database (1) wherever possible.2.To have tools specifically, but not exclusively, applicable to proteins prepared by 2-D gel electrophoresis.3.To have all tools available on the World Wide Web and freely available to the scientific community. [ABSTRACT FROM AUTHOR]

A federated two-dimensional (2-D) database is a database containing 2-D data that are accessible on the World Wide Web (WWW) and that are dynamically linked to other similar databases through active hypertext links over the Internet (1,2). Federated 2-D databases respect five rules that can be found at URL: http://www.expasy.ch/ch2d/fed-rules.html. [ABSTRACT FROM AUTHOR]

Many laboratories have identified proteins on two-dimensional (2-D) maps, and have built a master gel and a related 2-D database containing data specific to the identified proteins. Several of these databases have been made available on the World Wide Web (WWW) (1), and many more will undoubtedly follow. This chapter shows how to retrieve data remotely from these databases over the Internet. [ABSTRACT FROM AUTHOR]

Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) images, when used to analyze protein expression in large sets of biological samples, are best processed by computer. In this manner, one can detect and quantify even the faintest spots, quantitatively compare 2-D images among each other, or identify protein expression changes across various sets of 2-D images, such as disease vs control, or different stages in a toxicological study (1). [ABSTRACT FROM AUTHOR]