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Elucidating the properties of intrinsically disordered
proteins (IDPs) and unfolded and partially folded states of globular proteins
is challenging owing to their heterogeneous and dynamic nature. Protein
unfolding and misfolding is a key feature of a broad range of debilitating diseases,
whilst the conformational propensities of intrinsically disordered proteins can
play a significant role in modulating their activity, and the properties of
unfolded states of globular proteins modulates their stability and tendency to
aggregate. Ion mobility-mass spectrometry (IM-MS) is a powerful method for interrogating
these systems, however limits in resolution and the difficulty in probing the
energetics of interconversions amongst heterogeneous ensembles are major
issues. Herein, using a quadrupole/cyclic-IM/ time-of-flight MS instrument, we show
how the combination of precursor mass selection, mobility selection (IMn)
and collisional activation (CA) allows the elucidation of complicated gas-phase
dynamic behavior. The methodology employed is general and is demonstrated using
a classic model globular protein, cytochrome C, and an aggregation-prone IDP, amylin.
CA allows investigations of protein
conformational dynamics and unfolding in the gas-phase for heterogeneous
mixtures, whilst the additional precursor mass selection capability provides
high resolution and selectivity, facilitating more in-depth investigation.
Understanding protein dynamics in the gas-phase will allow greater insight into
protein behaviour and allow application of gas-phase techniques to clinically