Connecting Protein Millisecond Conformational Dynamics to Protein Thermal Stability

Area of Science:

• Biochemistry and Molecular Biology
• Protein Dynamics and Stability

Background:

• Protein folded state stability is essential for function but poorly understood in relation to sequence.
• Previous research focused on amino acid composition and couplings in single conformations, neglecting protein dynamics.

Purpose of the Study:

• To investigate the impact of alanine mutations in ubiquitin's C-terminal β-strand (β5) on protein dynamics and thermal stability.
• To explore the relationship between millisecond-timescale dynamics and protein stability.

Main Methods:

• Systematic analysis of alanine mutations in ubiquitin's β5 strand.
• Integration of spectroscopic and computational approaches.
• Characterization of conformational states and their interconversion.

Main Results:

• A negative correlation was observed between millisecond dynamics and thermal stability.
• Alanine substitutions in flexible C-terminal residues enhanced protein thermostability.
• The thermally unfolded state retained secondary structure but lacked β5 engagement, resembling the millisecond dynamics transition state.

Conclusions:

• Alanine mutations modulating folded state stability relative to the partially unfolded state affect both protein dynamics and stability.
• Conformational dynamics play a critical role in protein stability, with implications for protein engineering and design.