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Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.
Elution profile analysis of SDS-induced subcomplexes by quantitative mass spectrometry.
Mol. Cell. Proteomics 13, 1382-1391 (2014)
Analyzing the molecular architecture of native multiprotein complexes via biochemical methods has so far been difficult and error prone. Protein complex isolation by affinity purification can define the protein repertoire of a given complex, yet, it remains difficult to gain knowledge of its substructure or modular composition. Here, we introduce SDS concentration gradient induced decomposition of protein complexes coupled to quantitative mass spectrometry and in silico elution profile distance analysis. By applying this new method to a cellular transport module, the IFT/lebercilin complex, we demonstrate its ability to determine modular composition as well as sensitively detect known and novel complex components. We show that the IFT/lebercilin complex can be separated into at least five submodules, the IFT complex A, the IFT complex B, the 14-3-3 protein complex and the CTLH complex, as well as the dynein light chain complex. Furthermore, we identify the protein TULP3 as a potential new member of the IFT complex A and showed that several proteins, classified as IFT complex B-associated, are integral parts of this complex. To further demonstrate EPASIS general applicability, we analyzed the modular substructure of two additional complexes, that of B-RAF and of 14-3-3-epsilon. The results show, that EPASIS provides a robust as well as sensitive strategy to dissect the substructure of large multiprotein complexes in a highly time- as well as cost-effective manner.
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Publikationstyp
Artikel: Journalartikel
Dokumenttyp
Wissenschaftlicher Artikel
Schlagwörter
Statistical-model; Protein Complexes; Cilia; Accuracy; Ranbpm
ISSN (print) / ISBN
1535-9476
e-ISSN
1535-9484
Zeitschrift
Molecular and Cellular Proteomics
Quellenangaben
Band: 13,
Heft: 5,
Seiten: 1382-1391
Verlag
American Society for Biochemistry and Molecular Biology
Verlagsort
Bethesda
Nichtpatentliteratur
Publikationen
Begutachtungsstatus
Peer reviewed