Title
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Top-down mass spectrometry of intact membrane protein complexes reveals oligomeric state and sequence information in a single experiment
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Author
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Abstract
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Here we study the intact stoichiometry and top-down fragmentation behavior of three integral membrane proteins which were natively reconstituted into detergent micelles: the mechano-sensitive ion channel of large conductance (MscL), the Kirbac potassium channel and the p7 viroporin from the hepatitis C virus. By releasing the proteins under nondenaturing conditions inside the mass spectrometer, we obtained their oligomeric sizes. Increasing the ion activation (collision energy) causes unfolding and subsequent ejection of a highly charged monomer from the membrane protein complexes. Further increase of the ion activation then causes collision-induced dissociation (CID) of the ejected monomers, with fragments observed which were predominantly found to stem from membrane-embedded regions. These experiments show how in a single experiment, we can probe the relation between higher-order structure and protein sequence, by combining the native MS data with fragmentation obtained from top-down MS. |
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Language
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English
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Source (journal)
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Protein science. - Cambridge
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Publication
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Cambridge
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2015
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ISSN
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0961-8368
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DOI
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10.1002/PRO.2703
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Volume/pages
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24
:8
(2015)
, p. 1292-1300
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ISI
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000358176100012
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Pubmed ID
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25970171
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Full text (Publisher's DOI)
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Full text (open access)
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