Defining the Mandate of Proteomics in the Post-Genomics Era Workshop Report (2002) / Chapter Skim
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Workshop Report
Workshop Report
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After the presentations attendees were involved in individual breakout sessions on a variety of topics, including protein separation and identification protein structure and function metabolic pathways and post-translational modifications implementation: necessary policy and infrastructure conditions for collaboration platforms: emerging technologies computational methods and bioinfo~matics clinical proteomics
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CIark, "Towards a Total Human Protein Map". Indeed, Congress was considering a project called the "Human Protein Index" long before the Human Genome Project had been conceived.
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What should be the role of the private sector, particularly those companies large and small that have a major stake in exploiting the results of the venous genome projects and proteomics initiatives? How can all of these stakeholders cooperate most effectively while still maintaining proprietary information where appropriate?
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~ ~ fi : ~ ~ ~ ~ :~_ ~` : ~ ~ : ~- I, proteomics is not merely protein chemistry. Symposium chair and Dean of the University of Michigan College of Pharmacy, George Kenyon, commented, "Proteomics is not just a mass spectrum of a spot on a gel." Perhaps the most useful definition of proteomics for our purposes is the broadest: Proteomics represents the effort to establish the identities, quantities, structures, and biochemical and cellular functions of all proteins in an organism, organ, or organelle, and how these properties vary in space, time, or physiological state.
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and have used the broad definition given earlier to allow a wide-ranging discussion of goals, techniques, opportunities, and challenges. ~ A, ~ ~ a, Lessons Learned from the Human Genome Project Francis Collins, director of the National Human Genome Research Institute, spoke about lessons learned from the Human Genome Project that might be applicable to the discussion of a
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. He suggested that it was presumptuous to say that the Human Genome Project is already behind us.
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Collins said that the most important area for investment in proteomics right now is technology development so that we can move these methods in the direction of being able to tackle a mammalian proteome without facing enormous costs and problems with quality of the data. A number of resources for genomics research continue to be generated that may help inform a proteomics effort, including multiple coverage of certain genomes and more specifically: Multiple genomic sequences from mouse (6x coverage)
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It was noted that protein expression in a given cell varies from none to abundant. Historically, for practical reasons, the abundant proteins have been investigated most extensively; however, some of the rarely expressed proteins and proteins that appear only in disease states may be among the more interesting.
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Protein Separation After more than a decade of effort in gene sequencing, reliable estimates of the number of human genes is still a matter of disagreement, speculation, and debate. From the point of view of proteomics, just the detection or enumeration of the numbers of expressed proteins defies prediction based on our current understanding of human cell-type protein composition and its modulation by myriad undefined post-translational modifications.
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Denis Hochstrasser from the University of Geneva, a founder of GeneProt Inc., GeneBio SA, the Swiss Institute of Bioinformatics, and one of the pioneers in the identification of proteins in 2D gels, took the lead in dealing with the topic of protein separation. He stated at the outset that he wanted to play the role of "devil's advocate": to describe some of the excitement in proteomics but also to describe some of the difficulties.
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While many structural and metabolic gene products may be the same between all cells, as one reviewer pointed out, cell-specific proteins will be important for understanding function and disease. An afternoon breakout session, devoted to the topic of "protein separation and identification," was led by Julio Cells; Alain Van Dorsselear, Louis Pasteur University, CARS; and A
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still an experimental mainstay in the croteomics community can detect protein , ~ ~ ~ - , , · ~ · 1 · ~ ~~ · ~ ~ · 1 ~ · ~ ~ ^ ^ ~ ~ · concentrations as low as mlcromolar, a sensltlvlt-Y sutilclent to 1clent1ty ~ mu plasma proteins, not including modified forms. Under ideal conditions, including the sieving out of abundant proteins, mass spectrometer can extend sensitivity three orders of magnitude to the nanomolar level.
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The proteins are cleaved into peptides and then desorbed using a laser and finally delivered directly into the mass spectrometer. SOURCE: Courtesy of Denis Hochstrasser, GeneProt, Inc.
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Dr. Aebersold's group has developed a general approach to quantitative proteomics based on automated tandem mass spectrometry, stable isotope dilution theory, and a suite of bioinformatics tools for data analysis.
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are used to perform quantitative proteomic analyses based upon automated tandem mass spectrometry, stable isotope dilution theory, and a suite of bioinformatic tools for data analysis. Stable isotope signatures are introduced into proteins at specific sites via chemical reactions.
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In collaboration with Sciex (a manufacturer of mass spectrometers)
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In collaboration with Sciex (a manufacturer of mass spectrometers) , Ruedi Aebersold's group has developed a mass spectrometry system that allows one to quantify all the detected peptides first and then to selectively sequence only those that show an interesting quantitative behavior.
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For example, there exist · alternative splicing - which may affect as many as I/4 or more of the genes in a higher eukaryote and can alter biochemical function either drastically or subtly, producing truncated proteins and proteins with different compositions · post-translational modification, such as phosphorylation and glycosidation (which often occur on numerous sites on the same protein) · pre-enzymes made for secretion and pro-enzymes that are activated by cleavage · acylation and ubiquitination · non-enzymatic modifications like oxidation, so a given protein exists in the cell in different oxidized states.
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An interesting example of a chemical function genomics program was given by Thomas Leyh, co-chair of the "Structure Function" breakout session, who outlined an initiative intended to provide a functional genomics counterpart to the structural initiative already under way. The core of this multifaceted program, the subject of a 2001 National Institutes of Health workshop, is to perform large-scale mutagenesis and protein functional studies to create a database that assigns catalytic, ligand-binding, or other functions to the highly conserved, non-structural core residues for every protein family.
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The focus of her proteomics research is to use X-ray crystallography and NMR spectroscopy to determine the three-dimensional structures of proteins on a genome-wide scale. She is particularly interested in examining the extent to which protein structure can reveal protein function.
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Once again it was stressed that proteomics should be considered as a much broader field than would be apparent Dom early efforts, which have focused on cataloging levels of protein expression. Ideally it should encompass efforts to obtain complete functional descriptions for the gene products in a cell or organism.
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, ~ ,, _~_ ____ -- - ~-~ r~~~-~ ~~~~ a, A distantly related goal is the eventual application of proteomlcs to environmental, agncultural, and veterinary research, research areas that are far less developed than clinical applications. Thus, essentially all the applications discussed in the formal lectures and breakout sessions centered on clinical applications.
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Development of Diagnostics Participants of the "Clinical Aspects" breakout session on diagnostics discussed the fact that although the experimental platform used in clinical settings to detect protein markers will change rapidly in the coming years, the underlying principles regarding the stages of going from the discovery of protein markers to their use as diagnostic tools in a community setting will remain reasonably constant. Consequently the criteria used to judge the quality of a marker or markers as diagnostics in a clinical setting are different from those used to evaluate the quality of a marker in the basic science setting.
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Celis noted, but he stated they are not yet ready for the study of complex tissue samples. Scott Patterson, vice president of proteomics at Celera prefers the high-throughput approaches to clinical applications of proteomics research.
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Without effective and integrated databases to store and retrieve these data, and advanced computational methods such as pattern recognition and other machine learning approaches to analyze and interpret them, the full implications of these data will not be realized. A few years ago the typical biologist may have had little reason to turn to a computer for insights or information.
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The breakout group "Computational Methods and Bioinformatics," led by Kimmen Sjolander, University of California, Berkeley, and Dagmar Ringe, Brandeis University, discussed database issues. Database infrastructure and interface design For histoncal reasons most biological databases have been produced pnmarily by the biological community, while most computational tools have been produced by the mathematical and computational communities.
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that basically laid out the blueprint that became the Human Genome Project, and a wise blueprint, indeed," said Francis Collins.
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That is one of the joys of the whole thing." Protein Structural Initiative ~ September 2000 the NIGMS initiated the support of seven centers to begin work on developing an approach to structural genomics in order to reap the benefits of the multiple genome projects being undertaken worldwide. Two more centers were subsequently added in September 2001, forming what is now known as the Protein Structural Initiative.
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Interactions across disciplinary boundaries that were crucial for the genome project may be even more so for proteomics due to the variety of expertise needed. The group examined how this might be coordinated and made accessible through shared user facilities.
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Myers. "Collaboration to speed up that process is really a big driver here and driving it more than just the basic research kind of ideas." Conclusion The most useful definition of proteomics is likely to be the broadest: proteomics represents the effort to establish the identities, quantities, structures, and biochemical and cellular functions of all proteins in an organism, organ, or organelle, and how these properties vary in space, time, and physiological state.
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