Proteomics strategy for identifying candidate bioactive proteins in complex mixtures: application to the platelet releasate.

Hdl Handle:
http://hdl.handle.net/10147/107712
Title:
Proteomics strategy for identifying candidate bioactive proteins in complex mixtures: application to the platelet releasate.
Authors:
O'Connor, Roisin; Cryan, Lorna M; Wynne, Kieran; de Stefani, Andreas; Fitzgerald, Desmond; O'Brien, Colm; Cagney, Gerard
Affiliation:
School of Biomolecular and Biomedical Science, Conway Institute, University College Dublin, Dublin 4, Ireland.
Citation:
Proteomics strategy for identifying candidate bioactive proteins in complex mixtures: application to the platelet releasate. 2010, 2010:107859 J. Biomed. Biotechnol.
Journal:
Journal of biomedicine & biotechnology
Issue Date:
2010
URI:
http://hdl.handle.net/10147/107712
DOI:
10.1155/2010/107859
PubMed ID:
20368775
Abstract:
Proteomic approaches have proven powerful at identifying large numbers of proteins, but there are fewer reports of functional characterization of proteins in biological tissues. Here, we describe an experimental approach that fractionates proteins released from human platelets, linking bioassay activity to identity. We used consecutive orthogonal separation platforms to ensure sensitive detection: (a) ion-exchange of intact proteins, (b) SDS-PAGE separation of ion-exchange fractions and (c) HPLC separation of tryptic digests coupled to electrospray tandem mass spectrometry. Migration of THP-1 monocytes in response to complete or fractionated platelet releasate was assessed and located to just one of the forty-nine ion-exchange fractions. Over 300 proteins were identified in the releasate, with a wide range of annotated biophysical and biochemical properties, in particular platelet activation, adhesion, and wound healing. The presence of PEDF and involucrin, two proteins not previously reported in platelet releasate, was confirmed by western blotting. Proteins identified within the fraction with monocyte promigratory activity and not in other inactive fractions included vimentin, PEDF, and TIMP-1. We conclude that this analytical platform is effective for the characterization of complex bioactive samples.
Language:
en
MeSH:
Blood Platelets; Blood Proteins; Blotting, Western; Cell Movement; Chromatography, Ion Exchange; Complex Mixtures; Humans; Monocytes; Organ Specificity; Proteomics; Reproducibility of Results
ISSN:
1110-7251

Full metadata record

DC FieldValue Language
dc.contributor.authorO'Connor, Roisinen
dc.contributor.authorCryan, Lorna Men
dc.contributor.authorWynne, Kieranen
dc.contributor.authorde Stefani, Andreasen
dc.contributor.authorFitzgerald, Desmonden
dc.contributor.authorO'Brien, Colmen
dc.contributor.authorCagney, Gerarden
dc.date.accessioned2010-07-15T12:40:34Z-
dc.date.available2010-07-15T12:40:34Z-
dc.date.issued2010-
dc.identifier.citationProteomics strategy for identifying candidate bioactive proteins in complex mixtures: application to the platelet releasate. 2010, 2010:107859 J. Biomed. Biotechnol.en
dc.identifier.issn1110-7251-
dc.identifier.pmid20368775-
dc.identifier.doi10.1155/2010/107859-
dc.identifier.urihttp://hdl.handle.net/10147/107712-
dc.description.abstractProteomic approaches have proven powerful at identifying large numbers of proteins, but there are fewer reports of functional characterization of proteins in biological tissues. Here, we describe an experimental approach that fractionates proteins released from human platelets, linking bioassay activity to identity. We used consecutive orthogonal separation platforms to ensure sensitive detection: (a) ion-exchange of intact proteins, (b) SDS-PAGE separation of ion-exchange fractions and (c) HPLC separation of tryptic digests coupled to electrospray tandem mass spectrometry. Migration of THP-1 monocytes in response to complete or fractionated platelet releasate was assessed and located to just one of the forty-nine ion-exchange fractions. Over 300 proteins were identified in the releasate, with a wide range of annotated biophysical and biochemical properties, in particular platelet activation, adhesion, and wound healing. The presence of PEDF and involucrin, two proteins not previously reported in platelet releasate, was confirmed by western blotting. Proteins identified within the fraction with monocyte promigratory activity and not in other inactive fractions included vimentin, PEDF, and TIMP-1. We conclude that this analytical platform is effective for the characterization of complex bioactive samples.-
dc.language.isoenen
dc.subject.meshBlood Platelets-
dc.subject.meshBlood Proteins-
dc.subject.meshBlotting, Western-
dc.subject.meshCell Movement-
dc.subject.meshChromatography, Ion Exchange-
dc.subject.meshComplex Mixtures-
dc.subject.meshHumans-
dc.subject.meshMonocytes-
dc.subject.meshOrgan Specificity-
dc.subject.meshProteomics-
dc.subject.meshReproducibility of Results-
dc.titleProteomics strategy for identifying candidate bioactive proteins in complex mixtures: application to the platelet releasate.en
dc.contributor.departmentSchool of Biomolecular and Biomedical Science, Conway Institute, University College Dublin, Dublin 4, Ireland.en
dc.identifier.journalJournal of biomedicine & biotechnologyen

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