Metastability of Protein Solution Structures in the Absence of a Solvent: Rugged Energy Landscape and Glass-like...

Area of Science:

• Biochemistry
• Structural Biology
• Mass Spectrometry

Background:

• Native ion mobility-mass spectrometry (IM-MS) is a powerful tool for proteome structural screening.
• However, IM-MS characterizes protein structures in a solvent-free environment, raising questions about biological relevance.
• Understanding protein behavior in the absence of solvent is crucial for interpreting IM-MS data.

Purpose of the Study:

• To investigate the unfolding of ubiquitin in a solvent-free environment using novel computational and experimental IM-MS methods.
• To determine the structural integrity and dynamics of solvent-free ubiquitin.
• To compare the unfolding pathways of ubiquitin in solvent-free and solution states.

Main Methods:

• Utilized newly developed computational and experimental ion mobility/ion mobility/mass spectrometry (IM-IM/MS) techniques.
• Investigated the unfolding dynamics of ubiquitin under solvent-free conditions.
• Performed time-resolved IM-IM/MS measurements and molecular dynamics simulations.

Main Results:

• Solvent-free ubiquitin maintains a largely native fold, including the β-grasp motif and α-helix.
• The protein exhibits kinetically stable subpopulations reflecting solution structural heterogeneity.
• Unfolding rate constants in solvent-free conditions are 800-20,000 times smaller than in water, effectively halting unfolding.
• The unfolding pathway shares similarities with solution-phase pathways, indicating a conserved mechanism.

Conclusions:

• The energy landscape of solvent-free proteins is rugged, similar to glassy systems.
• Protein dynamics are significantly influenced by amino acid sequence.
• Solvent plays a facilitating role in protein motions rather than a controlling one.