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Molecular Weight Assessment of Proteins in Total Proteome Profiles using 1D-PAGE and LC/MS/MS

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dc.contributor.author Ahmad, Q Rushdy en_US
dc.contributor.author Nguyen, Dat H en_US
dc.contributor.author Wingerd, Mark A en_US
dc.contributor.author Church, George M en_US
dc.contributor.author Steffen, Martin A en_US
dc.date.accessioned 2012-01-11T00:42:55Z
dc.date.available 2012-01-11T00:42:55Z
dc.date.copyright 2005 en_US
dc.date.issued 2005-6-8 en_US
dc.identifier.citation Ahmad, Q Rushdy, Dat H Nguyen, Mark A Wingerd, George M Church, Martin A Steffen. "Molecular weight assessment of proteins in total proteome profiles using 1D-PAGE and LC/MS/MS" Proteome Science 3:6. (2005) en_US
dc.identifier.issn 1477-5956 en_US
dc.identifier.uri http://hdl.handle.net/2144/3063
dc.description.abstract BACKGROUND. The observed molecular weight of a protein on a 1D polyacrylamide gel can provide meaningful insight into its biological function. Differences between a protein's observed molecular weight and that predicted by its full length amino acid sequence can be the result of different types of post-translational events, such as alternative splicing (AS), endoproteolytic processing (EPP), and post-translational modifications (PTMs). The characterization of these events is one of the important goals of total proteome profiling (TPP). LC/MS/MS has emerged as one of the primary tools for TPP, but since this method identifies tryptic fragments of proteins, it has not generally been used for large-scale determination of the molecular weight of intact proteins in complex mixtures. RESULTS. We have developed a set of computational tools for extracting molecular weight information of intact proteins from total proteome profiles in a high throughput manner using 1D-PAGE and LC/MS/MS. We have applied this technology to the proteome profile of a human lymphoblastoid cell line under standard culture conditions. From a total of 1×107 cells, we identified 821 proteins by at least two tryptic peptides. Additionally, these 821 proteins are well-localized on the 1D-SDS gel. 656 proteins (80%) occur in gel slices in which the observed molecular weight of the protein is consistent with its predicted full-length sequence. A total of 165 proteins (20%) are observed to have molecular weights that differ from their predicted full-length sequence. We explore these molecular-weight differences based on existing protein annotation. CONCLUSION. We demonstrate that the determination of intact protein molecular weight can be achieved in a high-throughput manner using 1D-PAGE and LC/MS/MS. The ability to determine the molecular weight of intact proteins represents a further step in our ability to characterize gene expression at the protein level. The identification of 165 proteins whose observed molecular weight differs from the molecular weight of the predicted full-length sequence provides another entry point into the high-throughput characterization of protein modification. en_US
dc.description.sponsorship U.S.Department of Energy (Postdoctoral Fellowship in Computational Molecular Biology and Bioinformatics); Boston University (Whitaker Foundation Leadership Award) en_US
dc.language.iso en en_US
dc.publisher BioMed Central en_US
dc.rights Copyright 2005 Ahmad et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. en_US
dc.rights.uri http://creativecommons.org/licenses/by/2.0 en_US
dc.title Molecular Weight Assessment of Proteins in Total Proteome Profiles using 1D-PAGE and LC/MS/MS en_US
dc.type article en_US
dc.identifier.doi 10.1186/1477-5956-3-6 en_US
dc.identifier.pubmedid 15941491 en_US
dc.identifier.pmcid 1182394 en_US


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Copyright 2005 Ahmad et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Except where otherwise noted, this item's license is described as Copyright 2005 Ahmad et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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