HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Alert me to new issues of the journal Download to citation manager Glossary Citing Articles Citing Articles via Google Scholar Google Scholar Search for Related Content Social Bookmarking                      What's this?

Molecular & Cellular Proteomics 6:S37, 2007.
© 2007 by The American Society for Biochemistry and Molecular Biology, Inc.

---

Abstracts

Session 4

Interrogation of Cellular Signalling Pathways Using Protein Microarrays

J. van Oostrum

Novartis Institutes of BioMedical Research, Novartis AG, Basel, Switzerland

Molecular signalling pathways are frequently triggered by extracellular molecules binding receptors and activating relay systems inside cells, leading to processes that affect cellular behaviour and fate. For many genetic disorders a link between disease and signalling pathways have been established. Consequently, a systematic analysis of dynamic cellular networks provides an opportunity for pharmaceutical discovery, by taking into consideration the complex biological context of drug targets, rather than observing the targets in isolation. Such analyses are, perhaps, ideally suited for a systems biology approach that integrates experimental data with computational modeling with the aims of discovering and validating new drug targets and biomarkers, as well as predicting potential "off target" effects of drug candidates. Informing, calibrating and validating mathematical models with experimental data is a key component of an applied systems biology investigation and a number of genomic and proteomic techniques can be employed to generate these crucial data sets. These techniques range from LC-MS/MS based discovery of phosphopeptides to genome-wide cDNA and RNAi functional screens. Recently, we implemented and optimized a proteomics platform based on "reverse" protein arrays (RPA) that is particularly suitable to monitoring cell signalling events. These arrays are based on the principle that complex protein mixtures or proteomes (such as cell or tissue lysates) are spotted in an array format and probed with selected fluorescent antibodies in a multiplexed manner. To ensure high levels of sensitivity and signal to noise ratio of these RPAs, we are using planar waveguide technology. The advantage of the evanescent field fluorescence detection ensures that only analyte-bound fluorescent antibodies contribute to the signal. This method make it feasible to obtain quantitative and kinetic protein expression profiles and signaling information in a wide variety biospecimen.

4.2

Dynamic Interplay between GlcNAc and Phosphate on Regulatory Proteins

G. Hart

Johns Hopkins University School of Medicine, Baltimore, MD

O-GlcNAcylation is a dynamic modification of Ser(Thr) residues on signaling, transcription and cytoskeletal proteins in all metazoans. GlcNAcylation is very abundant and wide-spread, and often competes with phosphorylation, but also it directly plays an important role as a nutrient/stress sensor underlying glucose toxicity and diabetes. Studies during the last two decades have revealed serious limitations in both electrospray MS/MS and MALDI-TOF MS for the analysis of GlcNAcylation. However, recently, several chemico-enzymatic methods have been developed for the site mapping and quantification of O-GlcNAc. In addition, both FTMS with ECD and ion-traps with ETD are proving valuable for analyzing labile O-GlcNAc residues on proteins. We are currently using these methods to better understand both the global and specific interplay between GlcNAcylation and phosphorylation.

Supported by NIH grants HD13563, CA42486, DK61671, DK71280, and NIH contract N01-HV-28180. Dr. Hart receives a share of royalty received by the university on sales of the CTD 110.6 antibody. Terms of this arrangement are managed by JHU.

4.3

Mass Spectrometry as a Detector for Protein Ubiquitination

S. Gygi

Department of Cell Biology, Harvard Medical School, Boston MA

Of the major cell regulatory pathways, the biochemical complexity and functional diversity of protein ubiquitination make it an excellent candidate for large-scale proteomic studies. From the profiling of ubiquitin-protein conjugates by shotgun sequencing to the quantitative analysis of polyubiquitin chains, mass spectrometry is making critical advances in ubiquitin biology. In this lecture, we will describe a method for the detection and quantification of the levels of substrate protein, total ubiquitin, and each potential chain linkage. We demonstrate these measurements for conjugates formed by APC-catalyzed ubiquitination of cyclin B1.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This Article Full Text (PDF) Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Alert me to new issues of the journal Download to citation manager Glossary Citing Articles Citing Articles via Google Scholar Google Scholar Search for Related Content Social Bookmarking                      What's this?