Dissecting the Large Noncovalent Protein Complex GroEL with Surface-Induced Dissociation and Ion Mobility–Mass Spectrometry

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Abstract

Tandem mass spectrometry is a tool to dissect noncovalent protein complexes into smaller substructures for quaternary structure analysis. The commonly used activation method, collision induced dissociation (CID), often provides limited structural information from the typical dissociation pattern where unfolded monomers are ejected from the protein complex. In contrast, surface-induced dissociation (SID) has been shown to be very effective at dissociating protein complexes with less unfolding than CID. We present here SID of a large noncovalent tetradecamer protein, GroEL (801 kDa). A wide variety of products, including heptamers representative of the native topology, are released from the precursor upon SID, significantly different from the ubiquitous monomer ejection in CID. Enhanced dissociation into heptamers is observed when the charge states of the GroEL precursor are reduced by adding triethylammonium acetate into the spraying buffer. Ion mobility is utilized after SID to separate products overlapping in m/z to simplify the SID spectra. Compact heptamers from the charge-reduced tetradecamer are clearly distinguished from other overlapping species. SID can be very useful for quaternary structure studies of large noncovalent protein complexes, as manifested by the GroEL data where the tetradecamer dissociates into heptamers, reflecting the native topology of the complex.

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