Unfolding bovine α -lactalbumin with T-jump: Characterizing disordered intermediates via time-resolved x-ray solution scattering and molecular dynamics simulations

Darren J. Hsu, Denis Leshchev, Irina Kosheleva, Kevin L. Kohlstedt*, Lin X. Chen

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

The protein folding process often proceeds through partially folded transient states. Therefore, a structural understanding of these disordered states is crucial for developing mechanistic models of the folding process. Characterization of unfolded states remains challenging due to their disordered nature, and incorporating multiple methods is necessary. Combining the time-resolved x-ray solution scattering (TRXSS) signal with molecular dynamics (MD), we are able to characterize transient partially folded states of bovine α-lactalbumin, a model system widely used for investigation of molten globule states, during its unfolding triggered by a temperature jump. We track the unfolding process between 20 μs and 70 ms and demonstrate that it passes through three distinct kinetic states. The scattering signals associated with these transient species are then analyzed with TRXSS constrained MD simulations to produce protein structures that are compatible with the inputsignals. Without utilizing any experimentally extracted kinetic information, the constrained MD simulation successfully drove the protein to an intermediate molten globule state; signals for two later disordered states are refined to terminal unfolded states. From our examination of the structural characteristics of these disordered states, we discuss the implications disordered states have onthe folding process, especially on the folding pathway. Finally, we discuss the potential applications and limitations of this method.

Original languageEnglish (US)
Article number105101
JournalJournal of Chemical Physics
Volume154
Issue number10
DOIs
StatePublished - Mar 14 2021

ASJC Scopus subject areas

  • Physics and Astronomy(all)
  • Physical and Theoretical Chemistry

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