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Issue 9, 2011
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Identification of cytoplasmic and membrane-associated complexes in human embryonic stem cells using blue native PAGE

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Abstract

Human embryonic stem cells (hESCs) have great potential for use in developmental biology studies, functional genomics applications, drug screening, and regenerative medicine. A detailed understanding of the molecular mechanisms that are responsible for maintaining the undifferentiated and pluripotent nature of hESCs is essential for their effective therapeutic application. It has become evident that many complex cellular processes are carried out by assemblies of protein molecules (protein complexes). Blue native polyacrylamide gel electrophoresis (BN-PAGE) has been used to separate protein complexes from whole cell lysates. Using BN-PAGE, we resolved cytoplasmic and membrane-associated complexes from hESCs and characterised their composition, stoichiometry, and dynamics by denaturing SDS-PAGE. The reliability of the fractionation was examined by western blot analysis of membrane and cytosolic markers. MALDI TOF/TOF mass spectrometry identified 119 cytosolic and 69 membrane proteins from the BN-PAGE proteome maps. Potential protein complexes were validated by computational prediction of possible proteinprotein interactions using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database. Based on BN-PAGE gels and validation by databases, 82 heteromultimeric and 47 homomultimeric protein complexes have been found in hESCs. Resolving some of the protein complexes provided insight into the function of previously uncharacterised complexes in hESCs.

Graphical abstract: Identification of cytoplasmic and membrane-associated complexes in human embryonic stem cells using blue native PAGE

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Publication details

The article was received on 06 Apr 2011, accepted on 07 Jun 2011 and first published on 05 Jul 2011


Article type: Paper
DOI: 10.1039/C1MB05135K
Mol. BioSyst., 2011,7, 2688-2701

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    Identification of cytoplasmic and membrane-associated complexes in human embryonic stem cells using blue native PAGE

    F. Shekari, A. Taei, T. Pan, P. Wang, H. Baharvand and G. H. Salekdeh, Mol. BioSyst., 2011, 7, 2688
    DOI: 10.1039/C1MB05135K

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