Charge Mediated Compaction and Rearrangement of Gas-Phase Proteins: A Case Study Considering Two Proteins at Opposing Ends of the Structure-Disorder Continuum

Jacquelyn R. Jhingree, Bruno Bellina, Kamila J. Pacholarz, Perdita E. Barran

Charge reduction in the gas phase provides a direct means of manipulating protein charge state, and when coupled to ion mobility mass spectrometry (IM-MS), it is possible to monitor the effect of charge on protein conformation in the absence of solution. Use of the electron transfer reagent 1,3-dicyanobenzene, coupled with IM-MS, allows us to monitor the effect of charge reduction on the conformation of two proteins deliberately chosen from opposite sides of the order to disorder continuum: bovine pancreatic trypsin inhibitor (BPTI) and beta casein. The ordered BPTI presents compact conformers for each of three charge states accompanied by narrow collision cross-section distributions ((TW)CCSDN2→He). Upon reduction of BPTI, irrespective of precursor charge state, the (TW)CCSN2→He decreases to a similar distribution as found for the nESI generated ion of identical charge. The behavior of beta casein upon charge reduction is more complex. It presents over a wide charge state range (9-28), and intermediate charge states (13-18) have broad (TW)CCSDN2→He with multiple conformations, where both compaction and rearrangement are seen. Further, we see that the (TW)CCSDN2→He of the latter charge states are even affected by the presence of radical anions. Overall, we conclude that the flexible nature of some proteins result in broad conformational distributions comprised of many families, even for single charge states, and the barrier between different states can be easily overcome by an alteration of the net charge. Graphical Abstract ᅟ.