Refolding of Denatured Poly-ubiquitin in the Gas Phase
Heard, Stephanie C.
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For decades now, despite understanding that a protein’s structure plays a major role in its function, scientists still do not have a clear idea of how exactly proteins fold to adopt their native structures. Recently, ion mobility mass spectrometry (IM-MS) has emerged as a valuable analytical tool used to probe protein shape in the gas phase. In this work, multiply charged denatured poly-ubiquitin ions were charge reduced via cation to anion proton transfer reactions (CAPTR) and analyzed with a quadrupole/ion trap/ion mobility/time-offlight mass spectrometer. Arrival time distributions (ATDs) were analyzed to determine how the structure of the ions respond to changes in the charge state. Our results indicate that highly charged ions fold to more compact structures as charge is removed via CAPTR, in agreement with previous studies. However, we have also observed that product ion CCS is dependent on precursor selection, contrary to previous results acquired for mono-ubiquitin. Additionally, we have employed cumulative distribution functions to monitor protein folding. We observe that very high charge state precursor ions fold via similar pathways, while lower charge state precursor ions fold via unique pathways. These findings have implications for future studies in protein folding pathways.