Issue 20, 2015

Comparing equilibrium and kinetic protein unfolding using time-resolved electrospray-coupled ion mobility mass spectrometry

Abstract

Protein unfolding intermediates are thought to play a critical role in conformational pathogenesis, acting as a ‘gateway’ to inactivation or pathogenic aggregation. Unfolding intermediates have long been studied either by populating partially-folded species at equilibrium using incresingly denaturing conditions, or by transiently populating ‘kinetic’ intermediates under fully denaturing conditions using a time-resolved approach (e.g. stopped-flow fluorescence). However, it is not clear that the folding intermediates populated under equilibrium conditions are comparable to intermediates transiently populated in kinetic experiments. In this work, we combine time-resolved electrospray (TRESI) with travelling wave Ion Mobility Spectrometry (IMS) for the first time to directly compare equilibrium and kinetic unfolding intermediates of cytochrome c. Our results show a high degree of correlation between all species populated under these substantially different regimes.

Graphical abstract: Comparing equilibrium and kinetic protein unfolding using time-resolved electrospray-coupled ion mobility mass spectrometry

Article information

Article type
Paper
Submitted
28 ኤፕሪ 2015
Accepted
22 ጁን 2015
First published
22 ጁን 2015

Analyst, 2015,140, 6973-6979

Author version available

Comparing equilibrium and kinetic protein unfolding using time-resolved electrospray-coupled ion mobility mass spectrometry

P. Liuni, B. Deng and D. J. Wilson, Analyst, 2015, 140, 6973 DOI: 10.1039/C5AN00843C

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