Alternate Dissociation Pathways Identified in Charge-Reduced Protein Complex Ions

摘要

Tandem mass spectrometry (MS) of large protein com-nplexes has proven to be capable of assessing the stoichi-nometry, connectivity, and structural details of multiproteinnassemblies. While the utility of tandem MS is withoutnquestion, a deeper understanding of the mechanism ofnprotein complex dissociation will undoubtedly drive thentechnology into new areas of enhanced utility and infor-nmation content. We present here the systematic analysisnof the charge state dependent decay of the noncovalentlynassociated complex of human transthyretin, generated byncollision-induced dissociation (CID). A crown ether basedncharge reduction approach was applied to generate intactntransthyretin tetramers with charge states ranging fromn15 to 7 . These nine charge states were subsequentlynanalyzed by means of tandem MS and ion mobilitynspectrometry. Three different charge-dependent mecha-nnistic regimes were identified: (1) common asymmetricndissociation involving ejection of unfolded monomers, (2)nexpulsion of folded monomers from the intact tetramer,nand (3) release of C-terminal peptide fragments from thenintact complex. Taken together, the results presentednhighlight the potential of charge state modulation as anmethod for directing the course of gas-phase dissociationnand unfolding of protein complexes.