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Maxim B. Prigozhin, Yanxin Liu, Anna Jean Wirth, Shobhna Kapoor, Roland Winter, Klaus Schulten, and Martin Gruebele. Misplaced helix slows down ultrafast pressure-jump protein folding. Proceedings of the National Academy of Sciences, USA, 110:8087-8092, 2013.

PRIG2013 Using a newly developed microsecond pressure jump apparatus, we monitor the refolding kinetics of the helix-stabilized five helix bundle protein $\lambda$*YA from 3 $\mu$s to 5 ms after a 1200 bar pressure drop. In addition to a microsecond phase, we observe a ‘slow’ 1.4 ms phase during refolding to the native state. Unlike temperature denaturation, pressure denaturation produces a highly reversible helix-coil-rich state. The difference highlights the importance of denatured initial condition in folding experiments, and leads us to assign a compact non-native helical ‘trap’ as the reason for slower pressure jump- induced refolding. To complement the experiments, we performed over 50 $\mu$s of all-atom molecular dynamics pressure-drop refolding simulations with four different force fields. Two of the force fields yield compact non-native states with misplaced $\alpha$-helix content with a few $\mu$s of the pressure drop. Our overall conclusion from experiment and simulation is that the pressure denatured state of $\lambda$*YA contains mainly residual helix and little β-sheet; following a fast pressure drop, at least some $\lambda$*YA forms misplaced helical structure within microseconds. We hypothesize that non-native helix at helix-turn interfaces traps the protein in compact non-native conformations. These traps delay the folding of at least some of the population for 1.4 ms en route to the native state. Based on molecular dynamics, we predict specific mutations at the helix-turn interfaces that should speed up refolding from the pressure denatured state, if this hypothesis is correct.

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