Abstract
The structure and folding of a protein in solution depends on noncovalent interactions within the protein and those with surrounding ions and molecules. Decoupling these interactions in solution is challenging, which has hindered the development of accurate physics-based models for structure prediction. Investigations of proteins in the gas phase can be used to selectively decouple factors affecting the structures of proteins. Here, we use cation-to-anion proton-transfer reactions (CAPTR) to reduce the charge states of denatured ubiquitin ions in the gas phase, and ion mobility to probe their structures. In CAPTR, a precursor charge state is selected (P) and reacted with monoanions to generate charge-reduced product ions (C). Following each CAPTR event, denatured ubiquitin ions (13+ to 6+) yield products that rapidly isomerize to structures that have smaller collision cross sections (Ω). The Ω values of CAPTR product ions depend strongly on C and very weakly on P. Pre- and post-CAPTR activation was then used to probe the potential-energy surfaces of the precursor and product ions, respectively. Post-CAPTR activation showed that ions of different P fold differently and populate different regions of the potential-energy surface of that ion. Finally, pre-CAPTR activation showed that the structures of protein ions can be indirectly investigated using ion mobility of their CAPTR product ions, even for subtle structural differences that are not apparent from ion mobility characterization of the activated precursor ions. More generally, these results show that CAPTR strongly complements existing techniques for characterizing the structures and dynamics of biological molecules in the gas phase.