Intermediates and transition states in protein folding

Area of Science:

• Protein folding dynamics
• Biophysics
• Computational biology

Background:

• Protein folding often proceeds through an ensemble of intermediates.
• Apomyoglobin (apoMb) and lysozyme are model systems for studying protein folding pathways.
• The kinetic partitioning mechanism (KPM) describes folding when some molecules are trapped in intermediates.

Purpose of the Study:

• To dissect the complex role of intermediates in protein folding.
• To investigate factors influencing the cooperativity of intermediate formation in apoMb.
• To explore the implications of rugged energy landscapes and multiple folding routes.

Main Methods:

• Experimental data analysis of apomyoglobin (apoMb) and lysozyme.
• Theoretical concepts and simulations of simple off-lattice protein models.
• Kinetic and equilibrium studies under varying conditions (pH, anion concentration).

Main Results:

• Apomyoglobin exhibits a dominant kinetic intermediate (I) that also exists at equilibrium.
• The cooperativity of U<-->I transition in apoMb is sequence and anion concentration dependent.
• Protein folding can follow the kinetic partitioning mechanism (KPM), as seen in lysozyme, where pH influences fast folding fractions.
• Simulations differentiate equilibrium (on-pathway, revisitable) from kinetic (off-pathway, rarely revisited) intermediates.
• Kinetic intermediates are energetically higher than equilibrium intermediates.

Conclusions:

• Intermediates play a crucial role in protein folding, with distinct properties for equilibrium and kinetic types.
• Factors like pH, anion concentration, and mutations significantly impact folding pathways and intermediate populations.
• Rugged energy landscapes can lead to kinetic partitioning, trapping molecules in non-native states.
• The diversity of the denatured state ensemble influences the plasticity of the transition state ensemble (TSE).